U.S. patent application number 16/276326 was filed with the patent office on 2019-10-03 for 2-aryl- and 2-heteroaryl-substituted 2-pyridazin-3(2h)-one compounds as inhibitors of fgfr tyrosine kinases.
The applicant listed for this patent is Array BioPharma Inc.. Invention is credited to Steven W. Andrews, James F. Blake, Adam Cook, Indrani W. Gunawardana, Kevin W. Hunt, Andrew T. Metcalf, David Moreno, Li Ren, Tony P. Tang.
Application Number | 20190300511 16/276326 |
Document ID | / |
Family ID | 57233921 |
Filed Date | 2019-10-03 |
View All Diagrams
United States Patent
Application |
20190300511 |
Kind Code |
A1 |
Andrews; Steven W. ; et
al. |
October 3, 2019 |
2-ARYL- AND 2-HETEROARYL-SUBSTITUTED 2-PYRIDAZIN-3(2H)-ONE
COMPOUNDS AS INHIBITORS OF FGFR TYROSINE KINASES
Abstract
Provided herein are compounds of the general Formula I:
##STR00001## and stereoisomers and pharmaceutically acceptable
salts or solvates thereof, in which X, R.sup.1, R.sup.2, R.sup.3,
Ring A and z have the meanings given in the specification, which
are inhibitors of FGFR1, FGFR2, FGFR3 and/or FGFR4 and are useful
in the treatment and prevention of diseases which can be treated
with an FGFR inhibitor, including diseases or disorders mediated by
FGFR1, FGFR2, FGFR3 and/or FGFR4.
Inventors: |
Andrews; Steven W.;
(Boulder, CO) ; Blake; James F.; (Boulder, CO)
; Cook; Adam; (Boulder, CO) ; Gunawardana; Indrani
W.; (Boulder, CO) ; Hunt; Kevin W.; (Longmont,
CO) ; Metcalf; Andrew T.; (Boulder, CO) ;
Moreno; David; (Boulder, CO) ; Ren; Li;
(Boulder, CO) ; Tang; Tony P.; (Boulder,
CO) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Array BioPharma Inc. |
Boulder |
CO |
US |
|
|
Family ID: |
57233921 |
Appl. No.: |
16/276326 |
Filed: |
February 14, 2019 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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15333148 |
Oct 24, 2016 |
10208024 |
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16276326 |
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62245956 |
Oct 23, 2015 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07D 403/14 20130101;
C12Q 1/6886 20130101; A61K 31/5377 20130101; G01N 33/574 20130101;
C07D 413/14 20130101; C07D 401/14 20130101; A61K 45/06 20130101;
A61P 35/00 20180101; A61K 31/501 20130101 |
International
Class: |
C07D 403/14 20060101
C07D403/14; C07D 413/14 20060101 C07D413/14; C07D 401/14 20060101
C07D401/14; G01N 33/574 20060101 G01N033/574; A61K 31/501 20060101
A61K031/501; A61K 45/06 20060101 A61K045/06; A61K 31/5377 20060101
A61K031/5377; C12Q 1/6886 20060101 C12Q001/6886 |
Claims
1.-27. (canceled)
28. A method for treating cancer in a patient in need thereof, the
method comprising administering to the patient a therapeutically
effective amount of a compound of general Formula I: ##STR00234##
and pharmaceutically acceptable salts thereof, wherein: X is N or
CH; Ring A is a 5-membered heteroaryl ring having 1-2 ring nitrogen
atoms; z is 1, 2 or 3; each R.sup.1 is independently selected from
the group consisting of: (a) hydrogen; (b) C1-C6 alkyl optionally
substituted with 1-3 fluoros, (c) hydroxy(C1-C6 alkyl)- optionally
substituted with 1-3 fluoros, (d) dihydroxy(C1-C6 alkyl)-
optionally substituted with 1-3 fluoros, (e) cyano(C1-C6 alkyl)-,
(f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (g) (C1-C3 alkoxy)C1-C6 alkyl-
optionally substituted with 1-3 fluoros, (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl,
or (1-3C)alkoxy; (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, (k)
hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2, (l) halogen, and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--; hetCyc.sup.1 is a 4-7 membered
saturated heterocyclic ring having 1-2 ring heteroatoms
independently selected from N and O, wherein said heterocyclic ring
is optionally substituted with one or more substituents
independently selected from the group consisting of fluoro, HO,
C1-C6 alkyl (optionally substituted with 1-3 fluoros), (C1-C6
alkoxy)C1-C6 alkyl- (optionally substituted with 1-3 fluoros),
(C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6 alkyl)-,
R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--; hetCyc.sup.2 is a
7-10 membered heterospirocyclic ring having 1-2 ring heteroatoms
independently selected from N and O, wherein said heterospirocyclic
ring is optionally substituted with one or more substituents
independently selected from the group consisting of C1-C6 alkyl
(optionally substituted with 1-3 fluoros), (C1-C6 alkoxy)C1-C6
alkyl- (optionally substituted with 1-3 fluoros), (C3-C6
cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6 alkyl)-, R.sup.cR.sup.dN-
and (C1-C6 alkyl)C(.dbd.O)--; hetAr.sup.1 is a 6-membered
heteroaryl ring having 1-2 ring nitrogen atoms, wherein said ring
is optionally substituted with one or more substituents
independently selected from C1-C6 alkyl and halogen; R.sup.2 is
Ar.sup.1 or hetAr.sup.2; Ar.sup.1 is phenyl substituted with one or
more groups independently selected from halogen, cyano, C1-C3
alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3
alkyl)C(.dbd.O)NH--, (cyclopropyl)C(.dbd.O)NH-- and
(cyclopropyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl and
C1-C3 alkoxy portions is optionally substituted with 1-3 fluoros;
hetAr.sup.2 is a 6-10 membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said ring is optionally substituted with
one or more groups independently selected from halogen, C1-C3
alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3
alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and (C3-C4
cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl and
C1-C3 alkoxy portions is optionally substituted with 1-3 fluoros;
R.sup.3 is H, C1-C4 alkyl or (C3-C4)cycloalkyl; and R.sup.a,
R.sup.b, R.sup.c and R.sup.d are independently hydrogen or C1-C6
alkyl optionally substituted with F, OH or C1-C6 alkoxy.
29.-30. (canceled)
31. A method of treating an FGFR-associated cancer in a patient,
the method comprising: determining if the cancer in the patient is
an FGFR-associated cancer; and administering to a patient
determined to have an FGFR-associated cancer a therapeutically
effective amount of a compound of general Formula I: ##STR00235##
and pharmaceutically acceptable salts thereof, wherein: X is N or
CH; Ring A is a 5-membered heteroaryl ring having 1-2 ring nitrogen
atoms; z is 1, 2 or 3; each R.sup.1 is independently selected from
the group consisting of: (a) hydrogen; (b) C1-C6 alkyl optionally
substituted with 1-3 fluoros, (c) hydroxy(C1-C6 alkyl)- optionally
substituted with 1-3 fluoros, (d) dihydroxy(C1-C6 alkyl)-
optionally substituted with 1-3 fluoros, (e) cyano(C1-C6 alkyl)-,
(f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (g) (C1-C3 alkoxy)C1-C6 alkyl-
optionally substituted with 1-3 fluoros, (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl,
or (1-3C)alkoxy; (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, (k)
hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2, (l) halogen, and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--; hetCyc.sup.1 is a 4-7 membered
saturated heterocyclic ring having 1-2 ring heteroatoms
independently selected from N and O, wherein said heterocyclic ring
is optionally substituted with one or more substituents
independently selected from the group consisting of fluoro, HO,
C1-C6 alkyl (optionally substituted with 1-3 fluoros), (C1-C6
alkoxy)C1-C6 alkyl- (optionally substituted with 1-3 fluoros),
(C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6 alkyl)-,
R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--; hetCyc.sup.2 is a
7-10 membered heterospirocyclic ring having 1-2 ring heteroatoms
independently selected from N and O, wherein said heterospirocyclic
ring is optionally substituted with one or more substituents
independently selected from the group consisting of C1-C6 alkyl
(optionally substituted with 1-3 fluoros), (C1-C6 alkoxy)C1-C6
alkyl- (optionally substituted with 1-3 fluoros), (C3-C6
cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6 alkyl)-, R.sup.cR.sup.dN-
and (C1-C6 alkyl)C(.dbd.O)--; hetAr.sup.1 is a 6-membered
heteroaryl ring having 1-2 ring nitrogen atoms, wherein said ring
is optionally substituted with one or more substituents
independently selected from C1-C6 alkyl and halogen; R.sup.2 is
Ar.sup.1 or hetAr.sup.2; Ar.sup.1 is phenyl substituted with one or
more groups independently selected from halogen, cyano, C1-C3
alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3
alkyl)C(.dbd.O)NH--, (cyclopropyl)C(.dbd.O)NH-- and
(cyclopropyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl and
C1-C3 alkoxy portions is optionally substituted with 1-3 fluoros;
hetAr.sup.2 is a 6-10 membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said ring is optionally substituted with
one or more groups independently selected from halogen, C1-C3
alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3
alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and (C3-C4
cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl and
C1-C3 alkoxy portions is optionally substituted with 1-3 fluoros;
R.sup.3 is H, C1-C4 alkyl or (C3-C4)cycloalkyl; and R.sup.a,
R.sup.b, R.sup.c and R.sup.d are independently hydrogen or C1-C6
alkyl optionally substituted with F, OH or C1-C6 alkoxy.
32.-36. (canceled)
37. The method of claim 31, wherein the step of determining if the
cancer in the patient is an FGFR-associated cancer includes
performing an assay to detect dysregulation in a FGFR gene, a FGFR
protein, or expression or activity or level of any of the same in a
sample from the patient.
38.-42. (canceled)
43. The method of claim 37, wherein the dysregulation in a FGFR
gene, a FGFR protein, or expression or activity or level of any of
the same is one or more point mutations in the FGFR gene.
44.-51. (canceled)
52. The method of claim 37, wherein the dysregulation in a FGFR
gene, a FGFR protein, or expression or activity or level of any of
the same, is a chromosome translocation that results in the
expression of a FGFR fusion protein.
53.-55. (canceled)
56. The method of claim 31, wherein the FGFR-associated cancer is
selected from the group consisting of: urothelial carcinoma, breast
carcinoma or cancer, endometriod endometrial cancer or endometrial
cancer, ovarian carcinoma or cancer, brain cancer,
cholangiocarcinoma or intrahepatic cholangiocarinoma, gastric or
stomach cancer, gastrointestinal stromal tumors, lung cancer,
pancreatic cancer, prostate cancer, colorectal carcinoma or cancer,
rectal cancer, renal cell carcinoma, neuroendocrine carcinoma, head
and neck (squamous) carcinoma or head and neck adenoid cystic
carcinoma, skin cancer, leiomyosarcoma, sarcoma, osteosarcoma,
bladder cancer, rhabdomyosarcoma or embryonal rhabdosarcoma,
esophageal cancer, hepatocellular carcinoma or liver cancer,
salivary gland tumors, glioblatoma multiforme, myxoid
lipocarcinoma, oral cancer, thyroid cancer or carcinoma, anaplastic
thyroid carcinoma, adenoid cystic carcinoma, glioblastoma
multiforme, myeloproliferative disorder/hematological malignancy,
phosphaturic mesenchymal tumor, cervical cancer, biliary tract
cancer, gallbladder cancer, spermatocytic seminoma, seborrheic
keratosis, testicular cancer, fallopian tube carcinoma, carcinoma
of unknown primary, gastroesophageal junction carcinoma, anal
squamous cell carcinoma, hemangioendothelioma, uterine
carcinosarcoma or uterine cancer, carcinoid, peritoneal carcinoma,
adrenal carcinoma, bone cancer, peripheral nerve sheath tumor,
glioma or paraganglioma, rosette forming glioneural tumor,
lymphoepithelioma, mesothelioma, dysembryoplastic neuroepithelial
tumor, and dedifferentiated liposarcoma.
57.-163. (canceled)
164. The method of claim 28, wherein X is N.
165. The method of claim 28, wherein X is CH.
166. The method of claim 28, wherein R.sup.2 is Ar.sup.1.
167. The method of claim 28, wherein R.sup.2 is HetAr.sup.2.
168. The method of claim 28, wherein Ring A is pyrazolyl.
169. The method of claim 28, wherein R.sup.3 is hydrogen.
170. The method of claim 28, wherein the compound of general
Formula I is selected from any one of Examples 1-83 or a
pharmaceutically acceptable salt thereof.
171. The method of claim 28, wherein the cancer is selected from
the group consisting of: urothelial carcinoma, breast carcinoma or
cancer, endometriod endometrial cancer or endometrial cancer,
ovarian carcinoma or cancer, brain cancer, cholangiocarcinoma or
intrahepatic cholangiocarinoma, gastric or stomach cancer,
gastrointestinal stromal tumors, lung cancer, pancreatic cancer,
prostate cancer, colorectal carcinoma or cancer, rectal cancer,
renal cell carcinoma, neuroendocrine carcinoma, head and neck
(squamous) carcinoma or head and neck adenoid cystic carcinoma,
skin cancer, leiomyosarcoma, sarcoma, osteosarcoma, bladder cancer,
rhabdomyosarcoma or embryonal rhabdosarcoma, esophageal cancer,
hepatocellular carcinoma or liver cancer, salivary gland tumors,
glioblatoma multiforme, myxoid lipocarcinoma, oral cancer, thyroid
cancer or carcinoma, anaplastic thyroid carcinoma, adenoid cystic
carcinoma, glioblastoma multiforme, myeloproliferative
disorder/hematological malignancy, phosphaturic mesenchymal tumor,
cervical cancer, biliary tract cancer, gallbladder cancer,
spermatocytic seminoma, seborrheic keratosis, testicular cancer,
fallopian tube carcinoma, carcinoma of unknown primary,
gastroesophageal junction carcinoma, anal squamous cell carcinoma,
hemangioendothelioma, uterine carcinosarcoma or uterine cancer,
carcinoid, peritoneal carcinoma, adrenal carcinoma, bone cancer,
peripheral nerve sheath tumor, glioma or paraganglioma, rosette
forming glioneural tumor, lymphoepithelioma, mesothelioma,
dysembryoplastic neuroepithelial tumor, and dedifferentiated
liposarcoma.
172. The method of claim 31, wherein X is N.
173. The method of claim 31, wherein X is CH.
174. The method of claim 31, wherein R.sup.2 is Ar.sup.1.
175. The method of claim 31, wherein R.sup.2 is HetAr.sup.2.
176. The method of claim 31, wherein Ring A is pyrazolyl.
177. The method of claim 31, wherein R.sup.3 is hydrogen.
178. The method of claim 31, wherein the compound of general
Formula I is selected from any one of Examples 1-83 or a
pharmaceutically acceptable salt thereof.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application is a divisional of U.S. patent Ser. No.
15/333,148, filed Oct. 24, 2016, which claims the benefit of U.S.
Provisional Application No. 62/245,956, filed on Oct. 23, 2015,
which are incorporated herein by reference in their entirety.
BACKGROUND
[0002] The present disclosure relates to novel compounds which
exhibit inhibition of fibroblast growth factor receptor tyrosine
kinases (FGFRs), in particular FGFR1, FGFR2, FGFR3 and/or FGFR4,
pharmaceutical compositions comprising the compounds, to processes
for making the compounds, and the use of the compounds in therapy.
More particularly, it relates to 2-aryl- and
2-heteroaryl-substituted 2-pyridazin-3(2H)-one compounds useful in
the treatment or prevention of diseases which can be treated with
an FGFR inhibitor, including diseases mediated by FGFR tyrosine
kinases.
[0003] Fibroblast growth factors (FGFs) and their receptors (FGFRs)
regulate a wide range of physiologic cellular processes, such as
embryonic development, differentiation, proliferation, survival,
migration, and angiogenesis.
[0004] The FGF family comprises 18 secreted ligands (FGFs) which
are readily sequestered to the extracellular matrix by heparin
sulfate proteoglycans (HPSGs). For signal propagation, FGFs are
released from the extracellular matrix by proteases or specific
FGF-binding proteins, with the liberated FGFs subsequently binding
to a cell surface FGF-receptor (FGFR) in a ternary complex
consisting of FGF, FGFR and HPSG (Beenken, A., Nat. Rev. Drug
Discov. 2009; 8:235-253).
[0005] There are five FGFRs, of which four (FGFRs 1-4) are highly
conserved single-pass transmembrane tyrosine kinase receptors
(Eswarakumar, V. P., Cytokine Growth Factor Rev., 2005;
16:139-149). The binding of an FGF to an FGFR leads to receptor
dimerization and transphosphorylation of tyrosine kinase domains
(Dieci, M. V., et al., Cancer Discov. 2013; 3:264-279; Korc, N.,
and Friesel, R. E., Curr. Cancer Drug Targets 2009; 5:639-651).
Activation of downstream signaling occurs via the intracellular
receptor substrate FGFR substrate 2 (FRS2) and phospholipase
C.gamma. (PLC-.gamma.), leading to subsequent upregulation of
RAS/mitogen-activated protein kinase (MAPK) and phosphoinositide
3-kinase (PI3K)/AKT signaling pathways. Other pathways can be
activated, including STAT-dependent signaling (Turner, N., Grose,
R., Nat. Ref. Cancer 2010; 10:116-129; Brooks, N. S., et al., Clin
Cancer Res. 2012; 18:1855-1862; Dienstmann, R., et al., Ann. Oncol.
2014; 25:552-563).
[0006] FGFR signaling components are frequently altered in human
cancer, and several preclinical models have provided compelling
evidence for the oncogenic potential of aberrant FGFR signaling in
carcinogenesis, thereby validating FGFR signaling as an attractive
target for cancer treatment.
[0007] The mechanisms by which FGFR signaling is dysregulated and
drive cancer are better understood in recent years, and include
activating mutations, FGFR gene amplification, chromosomal
translocations, autocrine and paracrine signaling, and altered FGFR
splicing.
SUMMARY OF THE INVENTION
[0008] It has now been found that 2-aryl- and
2-heteroaryl-substituted 2-pyridazin-3(2H)-one compounds are
inhibitors of FGFR1, FGFR2, FGFR3 and/or FGFR4, which are useful in
the treatment or prevention of diseases which can be treated with
an inhibitor of FGFR1, FGFR2, FGFR3 and/or FGFR4, including
diseases mediated by FGFR1, FGFR2, FGFR3 and/or FGFR4.
[0009] Accordingly, provided herein is a compound of the general
Formula I:
##STR00002##
or pharmaceutically acceptable salt or solvate thereof, wherein X,
Ring A, z, R.sup.1, R.sup.2 and R.sup.3 are as defined herein.
[0010] Also provided herein is a pharmaceutical composition
comprising a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof, in admixture with a pharmaceutically
acceptable diluent or carrier.
[0011] Also provided herein is a method of inhibiting cell
proliferation, in vitro or in vivo, the method comprising
contacting a cell with an effective amount of a compound of Formula
I or a pharmaceutically acceptable salt or solvate thereof, or a
pharmaceutical composition thereof as defined herein.
[0012] Also provided herein is a method of treating an
FGFR-associated disease or disorder in a patient in need of such
treatment, the method comprising administering to the patient a
therapeutically effective amount of a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof, or a
pharmaceutical composition thereof as defined herein.
[0013] Also provided herein is a method of treating cancer and/or
inhibiting metastasis associated with a particular cancer in a
patient in need of such treatment, the method comprising
administering to the patient a therapeutically effective amount of
a compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof or a pharmaceutical composition thereof as defined
herein.
[0014] Also provided herein is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof, or a
pharmaceutical composition thereof as defined herein for use in
therapy.
[0015] Also provided herein is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof or a
pharmaceutical composition thereof as defined herein for use in the
treatment of cancer and/or inhibiting metastasis associated with a
particular cancer.
[0016] Also provided herein is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof for use in the
inhibition of FGFR1, FGFR2, FGFR3 and/or FGFR4.
[0017] Also provided herein is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof or a
pharmaceutical composition thereof as defined herein, for use in
the treatment of an FGFR-associated disease or disorder.
[0018] Also provided herein is the use of a compound of Formula I
or a pharmaceutically acceptable salt or solvate thereof, as
defined herein in the manufacture of a medicament for the treatment
of cancer and/or inhibiting metastasis associated with a particular
cancer.
[0019] Also provided herein is a use of a compound of Formula I or
a pharmaceutically acceptable salt or solvate thereof, as defined
herein in the manufacture of a medicament for the inhibition of
activity of FGFR1, FGFR2 FGFR3 and/or FGFR4.
[0020] Also provided herein is the use of a compound of Formula I
or a pharmaceutically acceptable salt or solvate thereof, as
defined herein, in the manufacture of a medicament for the
treatment of an FGFR-associated disease or disorder.
[0021] Also provided herein is a method for treating cancer in a
patient in need thereof, the method comprising (a) determining if
the cancer is associated with a dysregulation of an FGFR gene, a
fibroblast growth factor receptor, or expression or activity or
level of any of the same (e.g., an FGFR-associated cancer); and (b)
if the cancer is determined to be associated with a dysregulation
of an FGFR gene, a fibroblast growth factor receptor, or expression
or activity or level of any of the same (e.g., an FGFR-associated
cancer), administering to the patient a therapeutically effective
amount of a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof, or a pharmaceutical composition
thereof.
[0022] Also provided herein is a method for reversing or preventing
acquired resistance to an anticancer drug, comprising administering
a therapeutically effective amount of a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof, to a patient
at risk for developing or having acquired resistance to an
anticancer drug. In some embodiments, the patient is administered a
dose of the anticancer drug (e.g., at substantially the same time
as a dose of a compound of Formula I or a pharmaceutically
acceptable salt or solvate thereof is administered to the
patient).
[0023] Also provided herein is a method of delaying and/or
preventing development of cancer resistant to an anticancer drug in
an individual, comprising concomitantly administering to the
individual (a) an effective amount of a compound of Formula I and
(b) an effective amount of the anticancer drug.
[0024] Also provided herein is a method of treating an individual
with cancer who has increased likelihood of developing resistance
to an anticancer drug, comprising concomitantly administering to
the individual (a) an effective amount of a compound of Formula I
and (b) an effective amount of the anticancer drug.
[0025] Also provided herein is a method for treating a disease
involving angiogenesis and/or neovascularization, comprising
administering to a subject in need thereof, a therapeutically
effective amount of a compound of Formula I.
[0026] Also provided herein is a method for inhibiting angiogenesis
in a tumor, which comprises contacting the tumor with a compound of
Formula I.
[0027] Also provided herein is a pharmaceutical combination for
treating cancer in a patient in need thereof, which comprises (a) a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, (b) an additional therapeutic agent, and (c)
optionally at least one pharmaceutically acceptable carrier, for
simultaneous, separate or sequential use for the treatment of
cancer, wherein the amounts of the compound of Formula I or
pharmaceutically acceptable salt or solvate thereof and of the
additional therapeutic agent are together effective in treating the
cancer. Also provided herein is a pharmaceutical composition
comprising such a combination. Also provided herein is the use of
such a combination for the preparation of a medicament for the
treatment of cancer. Also provided herein is a commercial package
or product comprising such a combination as a combined preparation
for simultaneous, separate or sequential use; and to a method of
treatment of cancer a patient in need thereof.
[0028] Also provided herein is a process for preparing a compound
of Formula I or a pharmaceutically acceptable salt or solvate
thereof.
[0029] Also provided herein is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof obtained by a
process of preparing the compound as defined herein.
[0030] In some embodiments, a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof exhibits potent
and selective FGFR inhibition. In some embodiments, said inhibition
occurs with relative sparing of FGFR1 inhibition. In certain
embodiments, a compound of Formula I or a pharmaceutically
acceptable salt or solvate thereof exhibits a relatively high
potency for FGFR2 and FGFR3 (e.g., FGFR3, e.g., FGFR3-TACC3
fusion). In certain embodiments, a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof provides
dose-dependent inhibition of tumor growth in RT112/84 FGFR3-TACC3
xenografts. In certain embodiments, a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof exhibits
(independently) greater selectivity for FGFR2 and/or FGFR3 (e.g.,
FGFR3) as compared to FGFR1 (e.g., exhibits greater selectivity for
FGFR3 over FGFR1 in enzyme and cell-based assays e.g., exhibit
greater cytotoxicity for FGFR2/3 than FGFR1 mutant cells). See
Lewin, et al, Journal of Clinical Oncolgy, 2015, 22, 3372.
[0031] In some embodiments, administration of a compound of Formula
I or a pharmaceutically acceptable salt or solvate thereof results
in less hyperphosphatemia than administration of a pan-FGFR
inhibitor (e.g., a pan-FGFR inhibitor, which when compared with the
Formula I compounds described herein, exhibits less selectivity for
FGFR2 and/or FGFR3 (e.g., FGFR3) as compared to FGFR1; e.g., a
pan-FGFR inhibitor that is less sparing of FGFR1 inhibition than
the Formula I compounds described herein). In view of the foregoing
and while not wishing to be bound by theory, it is believed that a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof can provide greater dosing/regimen flexibility
and/or efficacy than, for example, a pan-FGFR inhibitor (e.g., a
pan-FGFR inhibitor, which when compared with the Formula I
compounds described herein, exhibits less selectivity for FGFR2
and/or FGFR3 (e.g., FGFR3) as compared to FGFR1; e.g., a pan-FGFR
inhibitor that is less sparing of FGFR1 inhibition than the Formula
I compounds described herein). By way of example, and as the
skilled person will appreciate, the compounds described herein can
be administered at higher doses and/or with increased frequencies,
thereby providing higher drug exposure/target coverage, and done so
with reduced risk of causing unwanted (e.g., abnormal) increases in
blood phosphate levels, which in some instances can necessitate
administration of phosphate binders and/or temporary (e.g., drug
holidays) or permanent cessation of therapy to allow phosphate
levels to return to normal.
[0032] Accordingly, also provided are methods of treating a
FGFR-associated cancer in a patient, which include: (a)
administering to a patient identified or diagnosed as having an
FGFR-associated cancer one or more doses of a first FGFR inhibitor
over a treatment period; (b) determining the level of phosphate in
a biological sample comprising blood, serum, or plasma obtained
from the patient after the treatment period; (c) selecting a
patient having an elevated level of phosphate in the biological
sample as compared to a reference level of phosphate; and (d)
ceasing administration of the first FGFR inhibitor and initiating
administration of a therapeutically effective amount of a compound
as described herein or a pharmaceutically acceptable salt or
solvate thereof, or a pharmaceutical composition containing the
same, to the selected patient. In certain embodiments, the
treatment period is at least 7 days. In other embodiments, the
treatment period is at least 21 days. In certain embodiments, the
first FGFR inhibitor is JNJ-42756493 or BGJ398. By way of example,
the first FGFR inhibitor can be JNJ-42756493 and a daily dose of 6
mg to 12 mg of the first FGFR inhibitor is administered to the
patient over the treatment period (e.g., 7 days). As another
example, the first FGFR inhibitor can be BGJ398 and a daily dose of
50 mg to 125 mg of the first FGFR inhibitor is administered to the
patient over the treatment period (e.g., 21 days). In certain
embodiments, the patient is administered a therapeutically
effective amount of a phosphate binder over the treatment period.
In certain embodiments, step (d) further comprises ceasing
administration of the phosphate binder to the selected patient. In
certain embodiments, step (d) further includes administering a
decreased dose of the phosphate binder to the selected patient
relative to the dose of the phosphate binder administered to the
patient over the treatment period. JNJ-42756493 (erdafitinib) is
also known as JNJ-493 and has the following systematic name,
N1-(3,5-dimethoxyphenyl)-N2-isopropyl-N1-(3-(1-methyl-1H-pyrazol-4-yl)qui-
noxalin-6-yl)ethane-1,2-diamine, and the following structure:
##STR00003##
[0033] BGJ398 (infigratinib) has the following systematic name,
3-(2,6-dichloro-3,5-dimethoxyphenyl)-1-(6-((4-(4-ethylpiperazin-1-yl)phen-
yl)amino)pyrimidin-4-yl)-1-methylurea, and the following chemical
structure:
##STR00004##
[0034] Also provided herein are methods of treating a
FGFR-associated cancer in a patient, the method comprising
administering a therapeutically effective dose of a compound as
described herein or a pharmaceutically acceptable salt or solvate
thereof, or a pharmaceutical composition containing the same to a
patient identified or diagnosed as having an FGFR-associated cancer
over a treatment period of at least 8 days, wherein the patient is
determined to have about the same or a decreased level of phosphate
in one or more biological sample(s) comprising blood, serum, or
plasma obtained from the patient over the treatment period as
compared to a reference level of phosphate.
[0035] Also provided herein are methods of treating a
FGFR-associated cancer in a patient, the method comprising
administering a therapeutically effective dose of a compound as
described herein or a pharmaceutically acceptable salt or solvate
thereof, or a pharmaceutical composition containing the same to a
patient identified or diagnosed as having an FGFR-associated cancer
over a treatment period, wherein the patient is not administered a
phosphate binder over the treatment period.
[0036] Also provided herein are methods of treating a
FGFR-associated cancer in a patient, the method comprising
administering a therapeutically effective dose of a compound as
described herein or a pharmaceutically acceptable salt or solvate
thereof, or a pharmaceutical composition containing the same to a
patient identified or diagnosed as having an FGFR-associated cancer
over a treatment period, wherein the patient is further
administered a low dose of a phosphate binder over the treatment
period.
[0037] Also provided herein are methods of treating a patient
having a FGFR-associated cancer, the method comprising
administering a therapeutically effective dose of a compound as
described herein or a pharmaceutically acceptable salt or solvate
thereof, or a pharmaceutical composition containing the same to a
patient identified or diagnosed as having an FGFR-associated cancer
over a treatment period, wherein the patient does not experience or
is less likely to experience one or more of soft tissue
calcification, stomatitis, dry mouth, nail changes, fatigue,
asthenia, anorexia, malaise, and muscle aches over the treatment
period.
[0038] Unless otherwise defined, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which this invention belongs. Methods
and materials are described herein for use in the present
invention; other, suitable methods and materials known in the art
can also be used. The materials, methods, and examples are
illustrative only and not intended to be limiting. All
publications, patent applications, patents, sequences, database
entries, and other references mentioned herein are incorporated by
reference in their entirety. In case of conflict, the present
specification, including definitions, will control.
[0039] Other features and advantages of the invention will be
apparent from the following detailed description, and from the
claims.
DETAILED DESCRIPTION OF THE INVENTION
[0040] Provided herein is a compound of the general Formula I:
##STR00005##
[0041] or a pharmaceutically acceptable salt or solvate thereof,
wherein:
[0042] X is N or CH;
[0043] Ring A is a 5-membered heteroaryl ring having 1-2 ring
nitrogen atoms;
[0044] z is 1, 2 or 3;
[0045] each R.sup.1 is independently selected from the group
consisting of:
[0046] (a) hydrogen,
[0047] (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros),
[0048] (c) hydroxy(C1-C6 alkyl)- (optionally substituted with 1-3
fluoros),
[0049] (d) dihydroxy(C1-C6 alkyl)- (optionally substituted with 1-3
fluoros),
[0050] (e) cyano(C1-C6 alkyl)-,
[0051] (f) R.sup.aR.sup.bN(C1-C6 alkyl)-,
[0052] (g) (C1-C3 alkoxy)C1-C6 alkyl- (optionally substituted with
1-3 fluoros),
[0053] (h) (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and
said cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy,
[0054] (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
[0055] (j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1,
[0056] (k) hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2,
[0057] (l) halogen, and
[0058] (m) hetCyc.sup.1C(.dbd.O)CH.sub.2--;
[0059] hetCyc.sup.1 is a 4-7 membered saturated heterocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterocyclic ring is optionally substituted with one
or more substituents independently selected from the group
consisting of fluoro, HO, C1-C6 alkyl (optionally substituted with
1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted
with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0060] hetCyc.sup.2 is a 7-10 membered heterospirocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterospirocyclic ring is optionally substituted with
one or more substituents independently selected from the group
consisting of C1-C6 alkyl (optionally substituted with 1-3
fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted with
1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0061] hetAr.sup.1 is a 6-membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said ring is optionally substituted with
one or more substituents independently selected from C1-C6 alkyl
and halogen;
[0062] R.sup.2 is Ar.sup.1 or hetAr.sup.2;
[0063] Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3 alkyl)C(.dbd.O)NH--,
(cyclopropyl)C(.dbd.O)NH-- and (cyclopropyl)NHC(.dbd.O)--, wherein
each of said C1-C3 alkyl and C1-C3 alkoxy portions is optionally
substituted with 1-3 fluoros;
[0064] hetAr.sup.2 is a 6-10 membered heteroaryl ring having 1-2
ring nitrogen atoms, wherein said heteroaryl ring is optionally
substituted with one or more groups independently selected from
halogen, C1-C3 alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--,
(C1-C3 alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and
(C3-C4 cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl
and C1-C3 alkoxy portions is optionally substituted with 1-3
fluoros;
[0065] R.sup.3 is hydrogen, C1-C4 alkyl or (C3-C4)cycloalkyl;
and
[0066] R.sup.a, R.sup.b, R.sup.c and R.sup.d are independently
hydrogen or C1-C6 alkyl optionally substituted with F, OH or C1-C6
alkoxy.
[0067] In some embodiments of general formula (I):
[0068] X is N or CH;
[0069] Ring A is a 5-membered heteroaryl ring having 1-2 ring
nitrogen atoms;
[0070] z is 1, 2 or 3;
[0071] each R.sup.1 is independently selected from the group
consisting of:
[0072] (a) hydrogen,
[0073] (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros),
[0074] (c) hydroxy(C1-C6 alkyl)- (optionally substituted with 1-3
fluoros),
[0075] (d) dihydroxy(C1-C6 alkyl)- (optionally substituted with 1-3
fluoros),
[0076] (e) cyano(C1-C6 alkyl)-,
[0077] (f) R.sup.aR.sup.bN(C1-C6 alkyl)-,
[0078] (g) (C1-C3 alkoxy)C1-C6 alkyl- (optionally substituted with
1-3 fluoros),
[0079] (h) (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and
said cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy,
[0080] (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
[0081] (j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1,
[0082] (k) hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2, and
[0083] (1) halogen;
[0084] hetCyc.sup.1 is a 4-7 membered saturated heterocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterocyclic ring is optionally substituted with one
or more substituents independently selected from the group
consisting of fluoro, HO, C1-C6 alkyl (optionally substituted with
1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted
with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0085] hetCyc.sup.2 is a 7-10 membered heterospirocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterospirocyclic ring is optionally substituted with
one or more substituents independently selected from the group
consisting of C1-C6 alkyl (optionally substituted with 1-3
fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted with
1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0086] hetAr.sup.1 is a 6-membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said ring is optionally substituted with
one or more substituents independently selected from C1-C6 alkyl
and halogen;
[0087] R.sup.2 is Ar.sup.1 or hetAr.sup.2;
[0088] Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3 alkyl)C(.dbd.O)NH--,
(cyclopropyl)C(.dbd.O)NH-- and (cyclopropyl)NHC(.dbd.O)--, wherein
each of said C1-C3 alkyl and C1-C3 alkoxy portions is optionally
substituted with 1-3 fluoros;
[0089] hetAr.sup.2 is a 6 membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said heteroaryl ring is optionally
substituted with one or more groups independently selected from
halogen, C1-C3 alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--,
(C1-C3 alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and
(C3-C4 cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl
and C1-C3 alkoxy portions is optionally substituted with 1-3
fluoros;
[0090] R.sup.3 is hydrogen, C1-C4 alkyl or (C3-C4)cycloalkyl;
and
[0091] R.sup.a, R.sup.b, R.sup.c and R.sup.d are independently
hydrogen or C1-C6 alkyl optionally substituted with F, OH or C1-C6
alkoxy.
[0092] For complex chemical names employed herein, the substituent
group is named before the group to which it attaches. For example,
methoxyethyl comprises an ethyl backbone with a methoxy
substituent.
[0093] The term "halogen" as used herein means --F (sometimes
referred to herein as "fluoro" or fluoros"), --Cl, --Br and
--I.
[0094] The terms "C1-C3 alkyl" and "C1-C6 alkyl" as used herein
refer to a monovalent, saturated linear or branched hydrocarbon
chains having from one to three and one to six carbon atoms,
respectively. Examples include, but are not limited to, methyl,
ethyl, 1-propyl, isopropyl, 1-butyl, isobutyl, sec-butyl,
tert-butyl, 2-methyl-2-propyl, pentyl, pentan-3-yl and hexyl.
[0095] The term "hydroxy(C1-C6 alkyl)-" as used herein refers to a
monovalent, saturated linear or branched hydrocarbon chain having
from one to six carbon atoms, wherein any one of the carbon atoms
is substituted with a hydroxy (--OH) group.
[0096] The terms "dihydroxy(C1-C6 alkyl)-" and "dihydroxy(C3-C6
alkyl)-" as used herein refers to a monovalent, saturated linear or
branched hydrocarbon chain having from one to six carbon atoms or
three to six carbon atoms, respectively, wherein any two of the
carbon atoms are each substituted with a hydroxy group, provided
that both hydroxy groups are not attached to the same carbon
atom.
[0097] The term "cyano(C1-C6 alkyl)-" as used herein refers to a
monovalent, saturated linear or branched hydrocarbon chain having
from one to six carbon atoms, wherein any one of the carbon atoms
is substituted with a cyano (--CN) group.
[0098] The terms "C1-C3 alkoxy" and "C1-C6 alkoxy" as used herein
refer to groups that have the formula, --OR, wherein R is "C1-C3
alkyl" and "C1-C6 alkyl", respectively, as defined herein.
Illustrative examples include, but are not limited to, methoxy,
ethoxy, propoxy, 2-propoxy, butoxy, tert-butoxy, pentyloxy, and
hexyloxy.
[0099] The term "(C1-C6 alkoxy)(C1-C6 alkyl)-" as used herein
refers to a monovalent, saturated linear or branched hydrocarbon
chain having from one to six carbon atoms, wherein any one of the
carbon atoms is substituted with a C1-C6 alkoxy group as defined
herein.
[0100] The term "C3-C6 cycloalkyl" refers to a monovalent,
monocyclic, saturated hydrocarbon ring having from three to six
ring atoms. Illustrative examples include, but are not limited to,
cyclopropyl and cyclobutyl.
[0101] The term "C3-C6 cycloalkoxy" as used herein refers to a
group having the formula, --OR', wherein R' is "C3-C6 cycloalkyl"
as defined herein.
[0102] The term "(C3-C6 cycloalkoxy)C1-C6 alkyl-" as used herein
refers to a monovalent, saturated linear or branched hydrocarbon
chain having from one to six carbon atoms, wherein any one of the
carbon atoms is substituted with a C3-C6 cycloalkoxy group as
defined herein.
[0103] The term "heterocyclic" refers to a saturated, monovalent,
monocyclic ring having the indicated number of total ring atoms, in
which at least one of the ring atoms is a heteroatom (e.g., N or
O).
[0104] The term "heterospirocyclic ring" as used herein refers to a
bicyclic, saturated, spiro-C-fused (i.e., the two rings share a
common carbon atom) heterocyclic ring system having from seven to
ten total ring atoms, wherein from one to two of the ring atoms is
a heteroatom independently selected from the group consisting of N
and O, provided that the heteroatoms are not adjacent to one
another. Each ring independently contains from 3 to 7 ring atoms,
and when two of the ring atoms are heteroatoms, each of the
heteroatoms can be present in the same ring, or each can be present
in a different ring. Examples include 7-oxa-4-azaspiro[2.5]octane,
2-azaspiro[3.3]heptane, 2,6-diazaspiro[3.3]heptane,
2,5-diazaspiro[3.4]octane, 2,6-diazaspiro[3.4]octane,
1,6-diazaspiro[3.4]octane, 1,7-diazaspiro[4.4]nonane,
2,7-diazaspiro[4.4]nonane, 2,7-diazaspiro[3.5]nonane,
2,6-diazaspiro[3.5]nonane, 2,5-diazaspiro[3.5]nonane,
1,7-diazaspiro[3.5]nonane, 1,6-diazaspiro[3.5]nonane,
2,8-diazaspiro[4.5]decane, 1,8-diazaspiro[4.5]decane,
1,7-diazaspiro[4.5]decane, 2,7-diazaspiro[4.5]decane,
2,6-diazaspiro[4.5]decane, 3,9-diazaspiro[5.5]undecane,
2,9-diazaspiro[5.5]undecane, 7-azaspiro[3.5]nonane,
6-oxa-2-azaspiro[3.4]octane, 2-oxa-7-azaspiro[4.4]nonane,
7-oxa-2-azaspiro[3.5]nonane and 7-oxa-2-azaspiro[4.5]decane. For
purposes of clarification, the chemical structures of two exemplary
heterospirocyclic rings are provided:
##STR00006##
[0105] The term "(C3-C4)cycloalkyl" as used herein refers
collectively to the cyclopropyl and cyclobutyl rings.
[0106] The term "heteroaryl" refers to an aromatic, monovalent or
divalent, monocyclic or bicyclic ring having the indicated number
of total ring atoms, in which at least one of the ring atoms is a
heteroatom (e.g., N or O).
[0107] The term "compound," as used herein is meant to include all
stereoisomers, geometric isomers, tautomers, and isotopically
enriched variants of the structures depicted. Compounds herein
identified by name or structure as one particular tautomeric form
are intended to include other tautomeric forms unless otherwise
specified.
[0108] The term "tautomer" as used herein refers to compounds whose
structures differ markedly in arrangement of atoms, but which exist
in easy and rapid equilibrium, and it is to be understood that
compounds of the present invention may be depicted as different
tautomers, and when compounds have tautomeric forms, all tautomeric
forms are intended to be within the scope of the invention, and the
naming of the compounds does not exclude any tautomer.
[0109] In certain embodiments of Formula I, X is N.
[0110] In certain embodiments of Formula I, X is CH.
[0111] In certain embodiments of Formula I, Ring A is pyrazolyl
optionally substituted with one to three R.sup.1 groups, wherein
each R.sup.1 group is independently selected from the group
consisting of (a) hydrogen, (b) C1-C6 alkyl (optionally substituted
with 1-3 fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally
substituted with 1-3 fluoros), (d) dihydroxy(C1-C6
alkyl)-(optionally substituted with 1-3 fluoros), (e) cyano(C1-C6
alkyl)-, (f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (g) (C1-C3 alkoxy)C1-C6
alkyl- (optionally substituted with 1-3 fluoros), (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, (k)
hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2, (1) halogen and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--. In certain embodiments, z is
1.
[0112] In certain embodiments of Formula I, Ring A is pyrazolyl
optionally substituted with one to three R.sup.1 groups, wherein
each R.sup.1 group is independently selected from the group
consisting of (a) hydrogen, (b) C1-C6 alkyl (optionally substituted
with 1-3 fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally
substituted with 1-3 fluoros), (e) cyano(C1-C6 alkyl)-, (g) (C1-C3
alkoxy)C1-C6 alkyl- (optionally substituted with 1-3 fluoros), (h)
(C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said
cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy, (i)
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3, (j)
hetCyc.sup.2(CH.sub.2).sub.p where p is 0 or 1, (1) halogen and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--. In certain embodiments, z is
1.
[0113] In certain embodiments of Formula I, R.sup.1 is hydrogen. In
one embodiment of Formula I, R.sup.1 is hydrogen and z is 1, 2 or
3.
[0114] In certain embodiments of Formula I, R.sup.1 is C1-C6 alkyl
optionally substituted with 1-3 fluoros. Non-limiting examples of
R.sup.1 include methyl, isopropyl, isobutyl, pentan-3-yl,
2,2-difluoroethyl, and 3,3,3-trifluoroethyl.
[0115] In certain embodiments of Formula I, R.sup.1 is C1-C6 alkyl
optionally substituted with 1-3 fluoros and z is 1, 2 or 3. In
certain embodiments, z is 1. In certain embodiments of Formula I,
Ring A is substituted with one to two R.sup.1 groups independently
selected from C1-C6 alkyl optionally substituted with 1-3 fluoros.
In certain embodiments of Formula I, Ring A is substituted with one
or two methyl groups. In certain embodiments of Formula I, Ring A
is substituted with two or three groups independently selected from
methyl and trifluoromethyl.
[0116] In certain embodiments of Formula I, R.sup.1 is
hydroxy(C1-C6 alkyl)- optionally substituted with 1-3 fluoros, and
z is 1, 2 or 3. In certain embodiments of Formula I, R.sup.1 is
hydroxy(C3-C6 alkyl)- optionally substituted with 1-3 fluoros. In
certain embodiments of Formula I, R.sup.1 is hydroxy(C1-C6 alkyl)
optionally substituted with 1-3 fluoros and z is 1. Non-limiting
examples of R.sup.1 include the structures:
##STR00007##
[0117] In certain embodiments of Formula I, R.sup.1 is
dihydroxy(C1-C6 alkyl)- optionally substituted with 1-3 fluoros,
and z is 1, 2 or 3. In certain embodiments of Formula I, R.sup.1 is
dihydroxy(C1-C6 alkyl)- optionally substituted with 1-3 fluoros and
z is 1. In certain embodiments, R.sup.1 is dihydroxy(C3-C6 alkyl)-.
Non-limiting examples of R.sup.1 include the structures:
##STR00008##
[0118] In certain embodiments of Formula I, R.sup.1 is cyano(C1-C6
alkyl)-, and z is 1, 2 or 3. In certain embodiments of Formula I,
R.sup.1 is cyano(C1-C6 alkyl)- and z is 1. A non-limiting example
of R.sup.1 includes the structure:
##STR00009##
[0119] In certain embodiments of Formula I, R.sup.1 is
R.sup.aR.sup.bN(C1-C6 alkyl)-, where R.sup.a and R.sup.b are
independently hydrogen or C1-C6 alkyl optionally substituted with
F, OH or C1-C6 alkoxy, and z is 1, 2 or 3. In certain embodiments
of Formula I, R.sup.1 is R.sup.aR.sup.bN(C1-C6 alkyl)- and z is 1.
Non-limiting examples of R.sup.1 include the structures:
##STR00010##
[0120] In certain embodiments of Formula I, R.sup.1 is (C1-C3
alkoxy)C1-C6 alkyl- optionally substituted with 1-3 fluoros, and z
is 1, 2 or 3. In certain embodiments of Formula I, R.sup.1 is
(C1-C3 alkoxy)C1-C6 alkyl- optionally substituted with 1-3 fluoros
and z is 1. Non-limiting examples of R.sup.1 include the
structures:
##STR00011##
[0121] In certain embodiments of Formula I, R.sup.1 is (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl,
or (1-3C)alkoxy, and z is 1, 2 or 3. In certain embodiments of
Formula I, R.sup.1 is (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n
is 0-3 and said cycloalkyl is optionally substituted with CN. In
certain embodiments, n is 0 or 1. In certain embodiments of Formula
I, R.sup.1 is (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- and z is 1.
Non-limiting examples of R.sup.1 include the structures:
##STR00012##
[0122] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.1(CH.sub.2).sub.m--, and z is 1, 2 or 3, where m is 0-3,
and hetCyc.sup.1 is a 4-7 membered saturated heterocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterocyclic ring is optionally substituted with one
or more substituents independently selected from the group
consisting of fluoro, HO, C1-C6 alkyl (optionally substituted with
1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl-(optionally substituted
with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--.
[0123] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.1(CH.sub.2).sub.m and z is 1, 2 or 3, where m is 0-3,
and hetCyc.sup.1 is azetidinyl, piperidinyl or morpholinyl
optionally substituted with one or more substituents independently
selected from the group consisting of fluoro, HO, C1-C6 alkyl
(optionally substituted with 1-3 fluoros), (C1-C6 alkoxy)C1-C6
alkyl- (optionally substituted with 1-3 fluoros), (C3-C6
cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6 alkyl), R.sup.cR.sup.dN-
and (C1-C6 alkyl)C(.dbd.O)--.
[0124] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.1(CH.sub.2).sub.m--, where m is 0-3, and hetCyc.sup.1 is
azetidinyl, piperidinyl or morpholinyl optionally substituted with
one or more substituents independently selected from the group
consisting of C1-C6 alkyl (optionally substituted with 1-3 fluoros)
and (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted with 1-3
fluoros). In certain embodiments, z is 1.
[0125] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.1(CH.sub.2).sub.m and z is 1. In one embodiment, m is 0,
1 or 2. In one embodiment, m is 0. In one embodiment, m is 1. In
one embodiment, m is 2.
[0126] Non-limiting examples of R.sup.1 when represented by
hetCyc.sup.1(CH.sub.2).sub.m-- include the structures:
##STR00013##
[0127] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and hetCyc.sup.2
is a 7-10 membered heterospirocyclic ring having 1-2 ring
heteroatoms independently selected from N and O, wherein said
heterospirocyclic ring is optionally substituted with one or more
substituents independently selected from the group consisting of
C1-C6 alkyl (optionally substituted with 1-3 fluoros), (C1-C6
alkoxy)C1-C6 alkyl- (optionally substituted with 1-3 fluoros),
(C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6 alkyl)-,
R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--, and z is 1, 2 or
3.
[0128] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and hetCyc.sup.2
is a 7-10 membered heterospirocyclic ring having 1-2 ring
heteroatoms independently selected from N and O, wherein said
heterospirocyclic ring is unsubstituted.
[0129] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.2(CH.sub.2).sub.p-- and z is 1.
[0130] Non-limiting examples when R.sup.1 is represented by
hetCyc.sup.2(CH.sub.2).sub.p-- include the structures:
##STR00014##
[0131] In certain embodiments of Formula I, R.sup.1 is
hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2 and hetAr.sup.1 is
a 6-membered heteroaryl ring having 1-2 ring nitrogen atoms,
wherein said ring is optionally substituted with one or more
substituents independently selected from C1-C6 alkyl and halogen,
and z is 1, 2 or 3.
[0132] In certain embodiments of Formula I, R.sup.1 is
hetAr.sup.1(CH.sub.2).sub.q-- and z is 1.
[0133] Non-limiting examples when R.sup.1 is represented by
hetAr.sup.1(CH.sub.2).sub.q-- include the structures:
##STR00015##
[0134] In certain embodiments of Formula I, R.sup.1 is halogen. In
certain embodiments when R.sup.1 is halogen, z is 1, 2 or 3.
Non-limiting examples of R.sup.1 include F, Cl and Br. In one
embodiment of Formula I, R.sup.1 is F and z is 1 or 2. In one
embodiment, z is 1.
[0135] In certain embodiments of Formula I, R.sup.1 is
hetCyc.sup.1C(.dbd.O)CH.sub.2--, and z is 1, 2 or 3, where m is
0-3, and hetCyc.sup.1 is a 4-7 membered saturated heterocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterocyclic ring is optionally substituted with one
or more substituents independently selected from the group
consisting of fluoro, HO, C1-C6 alkyl (optionally substituted with
1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl-(optionally substituted
with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--. In one
embodiment, hetCyc.sup.1 is piperidinyl or morpholinyl optionally
substituted with one or more substituents independently selected
from the group consisting of fluoro, HO, C1-C6 alkyl (optionally
substituted with 1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl-
(optionally substituted with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6
alkyl-, hydroxy(C1-C6 alkyl), R.sup.cR.sup.dN- and (C1-C6
alkyl)C(.dbd.O)--. In certain embodiments, hetCyc.sup.1 is
piperidinyl or morpholinyl optionally substituted with one or more
substituents independently selected from the group consisting of
C1-C6 alkyl (optionally substituted with 1-3 fluoros) and (C1-C6
alkoxy)C1-C6 alkyl- (optionally substituted with 1-3 fluoros). In
certain embodiments, z is 1. A non-limiting example of
hetCyc.sup.1C(.dbd.O)CH.sub.2-- is the structure:
##STR00016##
[0136] In certain embodiments of Formula I, Ring A is pyrazolyl
optionally substituted with one to three R.sup.1 groups (that is, z
is 1, 2 or 3), wherein R.sup.1 is selected from the group
consisting of (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros), (c) hydroxy(C1-C6 alkyl)-(optionally substituted with 1-3
fluoros), (e) cyano(C1-C6 alkyl)-, (f) R.sup.aR.sup.bN(C1-C6
alkyl)-, (h) (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3
and said cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy, (i)
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3, (j)
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--. In certain embodiments, z is 1
[0137] In certain embodiments of Formula I, Ring A is pyrazolyl
optionally substituted with two or three R.sup.1 groups (that is, z
is 2 or 3), wherein each R.sup.1 is independently selected from
hydrogen and C1-C6 alkyl (optionally substituted with 1-3
fluoros).
[0138] In certain embodiments of Formula I, z is 1 and Ring A is
pyrazolyl, which may be represented by the structure:
##STR00017##
[0139] wherein the wavy line indicates the point of attachment to
the 6-membered ring comprising X and the asterisk indicates the
point of attachment to R.sup.1, wherein R.sup.1 is selected from
(a) hydrogen, (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally substituted with
1-3 fluoros), (e) cyano(C1-C6 alkyl), (g) (C1-C3 alkoxy)C1-C6
alkyl- (optionally substituted with 1-3 fluoros), (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p where p is 0 or 1, (1) halogen and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--. In one embodiment, R.sup.1 is
selected from (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally substituted with
1-3 fluoros), (e) cyano(C1-C6 alkyl)-, (f) R.sup.aR.sup.bN(C1-C6
alkyl)-, (h) (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3
and said cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy, (i)
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3, (j)
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--.
[0140] In certain embodiments of Formula I, Ring A is pyrazolyl and
z is 1, wherein Ring A and R.sup.1 together may be represented by
the structure:
##STR00018##
[0141] wherein the wavy line indicates the point of attachment to
the 6-membered ring comprising X, wherein z is 1 and R.sup.1 is
selected from (a) hydrogen, (b) C1-C6 alkyl (optionally substituted
with 1-3 fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally
substituted with 1-3 fluoros), (e) cyano(C1-C6 alkyl)-, (g) (C1-C3
alkoxy)C1-C6 alkyl- (optionally substituted with 1-3 fluoros), (h)
(C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said
cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy, (i)
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3, (j)
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, (1) halogen and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--. In one embodiment, R.sup.1 is
selected from (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally substituted with
1-3 fluoros), (e) cyano(C1-C6 alkyl)-, (f) R.sup.aR.sup.bN(C1-C6
alkyl)-, (h) (C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3
and said cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy, (i)
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--.
[0142] In certain embodiments of Formula I, R.sup.2 is Ar.sup.1,
where Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3 alkyl)C(.dbd.O)NH--,
(cyclopropyl)C(.dbd.O)NH-- and (cyclopropyl)NHC(.dbd.O)--, wherein
each of said C1-C3 alkyl and C1-C3 alkoxy portions are optionally
substituted with 1-3 fluoros.
[0143] In certain embodiments of Formula I, R.sup.2 is Ar.sup.1,
where Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy and (C1-C3 alkyl)NHC(.dbd.O)--, wherein each of said C1-C3
alkyl and C1-C3 alkoxy portions is optionally substituted with 1-3
fluoros.
[0144] Non-limiting examples of R.sup.2 when represented by
Ar.sup.1 include the structures:
##STR00019## ##STR00020##
[0145] In certain embodiments of Formula I, R.sup.2 is hetAr.sup.2,
where hetAr.sup.2 is a 6-10 membered heteroaryl ring having 1-2
ring nitrogen atoms, wherein said ring is optionally substituted
with one or more groups independently selected from halogen, C1-C3
alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3
alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and (C3-C4
cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl and
C1-C3 alkoxy portions is optionally substituted with 1-3
fluoros.
[0146] In certain embodiments of Formula I, R.sup.2 is hetAr.sup.2,
where hetAr.sup.2 is a 6-10 membered heteroaryl ring having 1-2
ring nitrogen atoms, wherein said ring is optionally substituted
with one or more groups independently selected from C1-C3 alkyl,
C1-C3 alkoxy and (C1-C3 alkyl)NHC(.dbd.O)--, wherein each of said
C1-C3 alkyl and C1-C3 alkoxy portions is optionally substituted
with 1-3 fluoros.
[0147] Non-limiting examples of hetAr.sup.2 include:
##STR00021##
[0148] In certain embodiments of Formula I, R.sup.3 is H.
[0149] In certain embodiments of Formula I, R.sup.3 is C1-C4 alkyl.
Non-limiting examples include methyl, ethyl, propyl, isopropyl and
isobutyl.
[0150] In certain embodiments of Formula I, R.sup.3 is
(C3-C4)cycloalkyl. In certain embodiments of Formula I, R.sup.3 is
cyclopropyl. In certain embodiments of Formula I, R.sup.3 is
cyclobutyl.
[0151] Compounds of Formula I include compounds of Formula I-A,
wherein:
[0152] X is N;
[0153] z is 1;
[0154] Ring A is
##STR00022##
[0155] wherein the wavy line indicates the point of attachment to
the 6-membered ring comprising X and the asterisk indicates the
point of attachment to R.sup.1;
[0156] each R.sup.1 is independently selected from the group
consisting of (a) hydrogen, (b) C1-C6 alkyl (optionally substituted
with 1-3 fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally
substituted with 1-3 fluoros), (d) dihydroxy(C1-C6 alkyl)-
(optionally substituted with 1-3 fluoros), (e) cyano(C1-C6 alkyl)-,
(f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (g) (C1-C3 alkoxy)C1-C6
alkyl-(optionally substituted with 1-3 fluoros), (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, (k)
hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2, (l) halogen and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--;
[0157] hetCyc.sup.1 is a 4-7 membered saturated heterocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterocyclic ring is optionally substituted with one
or more substituents independently selected from the group
consisting of fluoro, HO, C1-C6 alkyl (optionally substituted with
1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted
with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0158] hetCyc.sup.2 is a 7-10 membered heterospirocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterospirocyclic ring is optionally substituted with
one or more substituents independently selected from the group
consisting of C1-C6 alkyl (optionally substituted with 1-3
fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted with
1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0159] hetAr.sup.1 is a 6-membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said ring is optionally substituted with
one or more substituents independently selected from C1-C6 alkyl
and halogen;
[0160] R.sup.2 is Ar.sup.1 or hetAr.sup.2;
[0161] Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3 alkyl)C(.dbd.O)NH--,
(cyclopropyl)C(.dbd.O)NH-- and (cyclopropyl)NHC(.dbd.O)--, wherein
each of said C1-C3 alkyl and C1-C3 alkoxy portions is optionally
substituted with 1-3 fluoros;
[0162] hetAr.sup.2 is a 6-10 membered heteroaryl ring having 1-2
ring nitrogen atoms, wherein said heteroaryl ring is optionally
substituted with one or more groups independently selected from
halogen, C1-C3 alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--,
(C1-C3 alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and
(C3-C4 cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl
and C1-C3 alkoxy portions is optionally substituted with 1-3
fluoros;
[0163] R.sup.3 is hydrogen, C1-C4 alkyl or (C3-C4)cycloalkyl;
and
[0164] R.sup.a, R.sup.b, R.sup.c and R.sup.d are independently
hydrogen or C1-C6 alkyl optionally substituted with F, OH or C1-C6
alkoxy.
[0165] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is as defined for Formula I-A.
[0166] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--.
[0167] In one embodiment of Formula I-A, R.sup.3 is hydrogen.
[0168] In one embodiment of Formula I-A, R.sup.3 is C1-C4
alkyl.
[0169] In one embodiment of Formula I-A, R.sup.3 is
(C3-C4)cycloalkyl.
[0170] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--, and
R.sup.3 is hydrogen.
[0171] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--, and
R.sup.3 is C1-C4 alkyl.
[0172] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--, and
R.sup.3 is (C3-C4)cycloalkyl.
[0173] In one embodiment of Formula I-A, R.sup.1 is selected from
(b) C1-C6 alkyl (optionally substituted with 1-3 fluoros), (c)
hydroxy(C1-C6 alkyl)- (optionally substituted with 1-3 fluoros),
(e) cyano(C1-C6 alkyl)-, (f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (h)
(C3-C6 cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said
cycloalkyl is optionally substituted with CN, OH,
R.sup.aR.sup.bN--, (1-3C)alkyl or (1-3C)alkoxy, (i)
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3, (j)
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--; and hetCyc.sup.1, R.sup.a, and
R.sup.b are as defined for Formula I-A.
[0174] In one embodiment of Formula I-A, R.sup.1 is C1-C6 alkyl
(optionally substituted with 1-3 fluoros).
[0175] In one embodiment of Formula I-A, R.sup.1 is hydroxy(C1-C6
alkyl)- (optionally substituted with 1-3 fluoros).
[0176] In one embodiment of Formula I-A, R.sup.1 is cyano(C1-C6
alkyl)-.
[0177] In one embodiment of Formula I-A, R.sup.1 is
R.sup.aR.sup.bN(C1-C6 alkyl)-.
[0178] In one embodiment of Formula I-A, R.sup.1 is (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy,
[0179] In one embodiment of Formula I-A, R.sup.1 is
hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3. In one embodiment, m
is 0, 1 or 2. In one embodiment, m is 0. In one embodiment, m is 1.
In one embodiment, m is 2.
[0180] In one embodiment of Formula I-A, R.sup.1 is
hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1.
[0181] In one embodiment of Formula I-A, R.sup.1 is
hetCyc.sup.1C(.dbd.O)CH.sub.2--.
[0182] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; R.sup.1 is selected from (b) C1-C6 alkyl
(optionally substituted with 1-3 fluoros), (c) hydroxy(C1-C6
alkyl)- (optionally substituted with 1-3 fluoros), (e) cyano(C1-C6
alkyl)-, (f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--; and hetCyc.sup.1, R.sup.a, and
R.sup.b are as defined for Formula I-A. In one embodiment, m is 0,
1 or 2. In one embodiment, m is 0. In one embodiment, m is 1. In
one embodiment, m is 2.
[0183] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is C1-C4 alkyl; R.sup.1 is selected from (b) C1-C6 alkyl
(optionally substituted with 1-3 fluoros), (c) hydroxy(C1-C6
alkyl)- (optionally substituted with 1-3 fluoros), (e) cyano(C1-C6
alkyl)-, (f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--; and hetCyc.sup.1, R.sup.a, and
R.sup.b are as defined for Formula I-A. In one embodiment, m is 0,
1 or 2. In one embodiment, m is 0. In one embodiment, m is 1. In
one embodiment, m is 2.
[0184] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is (C3-C4)cycloalkyl; R.sup.1 is selected from (b) C1-C6
alkyl (optionally substituted with 1-3 fluoros), (c) hydroxy(C1-C6
alkyl)- (optionally substituted with 1-3 fluoros), (e) cyano(C1-C6
alkyl)-, (f) R.sup.aR.sup.bN(C1-C6 alkyl)-, (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, and (m)
hetCyc.sup.1C(.dbd.O)CH.sub.2--; and hetCyc.sup.1, R.sup.a, and
R.sup.b are as defined for Formula I-A. In one embodiment, m is 0,
1 or 2. In one embodiment, m is 0. In one embodiment, m is 1. In
one embodiment, m is 2.
[0185] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; and R.sup.1 is C1-C6 alkyl (optionally
substituted with 1-3 fluoros).
[0186] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkyl (optionally
substituted with 1-3 fluoros), and C1-C3 alkoxy (optionally
substituted with 1-3 fluoros); R.sup.3 is hydrogen; and R.sup.1 is
hydroxy(C1-C6 alkyl)- (optionally substituted with 1-3
fluoros).
[0187] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkyl (optionally
substituted with 1-3 fluoros), and C1-C3 alkoxy (optionally
substituted with 1-3 fluoros); R.sup.3 is hydrogen; and R.sup.1 is
cyano(C1-C6 alkyl)-.
[0188] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; and R.sup.1 is R.sup.aR.sup.bN(C1-C6 alkyl)-
where R.sup.a, and R.sup.b are as defined for Formula I-A.
[0189] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; and R.sup.1 is (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy. In one embodiment said cycloalkyl is
unsubstituted. In one embodiment, said cycloalkyl is substituted
with CN. In one embodiment, n is 0. In one embodiment n is 1.
[0190] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; R.sup.1 is hetCyc.sup.1(CH.sub.2).sub.m--
where m is 0-3; and hetCyc.sup.1 is as defined for Formula I-A. In
one embodiment, m is 0, 1 or 2. In one embodiment, m is 0. In one
embodiment, m is 1. In one embodiment, m is 2.
[0191] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; and R.sup.1 is hetCyc.sup.2(CH.sub.2).sub.p--
where p is 0 or 1, and hetCyc.sup.2 is as defined for Formula I-A.
In one embodiment, p is 1. On one embodiment,
hetCyc.sup.2(CH.sub.2).sub.p-- is
##STR00023##
[0192] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is hydrogen; and R.sup.1 is hetCyc.sup.1C(.dbd.O)CH.sub.2--
where hetCyc.sup.1 is as defined for Formula I-A. In one
embodiment, hetCyc.sup.1 is piperidinyl. In one embodiment,
hetCyc.sup.1C(.dbd.O)CH.sub.2-- is
##STR00024##
[0193] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is C1-C4 alkyl; and R.sup.1 is C1-C6 alkyl (optionally
substituted with 1-3 fluoros).
[0194] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is C1-C4 alkyl; and R.sup.1 is R.sup.aR.sup.bN(C1-C6
alkyl)-, wherein R.sup.a and R.sup.b are as defined for Formula
I-A. In one embodiment, R.sup.a and R.sup.b are independently
selected from C1-C6 alkyl.
[0195] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1, where
Ar.sup.1 is phenyl optionally substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl (optionally
substituted with 1-3 fluoros), C1-C3 alkoxy (optionally substituted
with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--; R.sup.3 is C1-C4
alkyl; and R.sup.1 is hetCyc.sup.1(CH.sub.2).sub.m-- where m is
0-3, and hetCyc.sup.1 is as defined for Formula I-A.
[0196] In one embodiment of Formula I-A, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, cyano, C1-C3 alkyl
(optionally substituted with 1-3 fluoros), C1-C3 alkoxy (optionally
substituted with 1-3 fluoros) and (C1-C3 alkyl)NHC(.dbd.O)--;
R.sup.3 is C3-C4 cycloalkyl; and R.sup.1 is C1-C6 alkyl (optionally
substituted with 1-3 fluoros).
[0197] Compounds of Formula I include compounds of Formula I-B,
wherein:
[0198] X is N;
[0199] z is 2 or 3;
[0200] Ring A is pyrazolyl;
[0201] each R.sup.1 is independently selected from the group
consisting of (a) hydrogen and (b) C1-C6 alkyl (optionally
substituted with 1-3 fluoros);
[0202] R.sup.2 is Ar.sup.1;
[0203] Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3 alkyl)C(.dbd.O)NH--,
(cyclopropyl)C(.dbd.O)NH-- and (cyclopropyl)NHC(.dbd.O)--, wherein
each of said C1-C3 alkyl and C1-C3 alkoxy portions is optionally
substituted with 1-3 fluoros; and
[0204] R.sup.3 is hydrogen, C1-C4 alkyl or (C3-C4)cycloalkyl.
[0205] In one embodiment of Formula I-B, Ar.sup.1 is phenyl
substituted with one or more groups independently selected from
halogen and C1-C3 alkoxy.
[0206] In one embodiment of Formula I-B, R.sup.3 is hydrogen.
[0207] In one embodiment of Formula I-B, Ar.sup.1 is phenyl
substituted with one or more groups independently selected from
halogen and C1-C3 alkoxy, and R.sup.3 is hydrogen.
[0208] Compounds of Formula I include compounds of Formula I-C,
wherein:
[0209] X is CH;
[0210] z is 1;
[0211] Ring A is
##STR00025##
[0212] wherein the wavy line indicates the point of attachment to
the 6-membered ring comprising X and the asterisk indicates the
point of attachment to R.sup.1;
[0213] R.sup.1 is selected from the group consisting of (a)
hydrogen, (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros), (c) hydroxy(C1-C6 alkyl)- (optionally substituted with
1-3 fluoros), (d) dihydroxy(C1-C6 alkyl)- (optionally substituted
with 1-3 fluoros), (e) cyano(C1-C6 alkyl)-, (f)
R.sup.aR.sup.bN(C1-C6 alkyl)-, (g) (C1-C3 alkoxy)C1-C6 alkyl-
(optionally substituted with 1-3 fluoros), (h) (C3-C6
cycloalkyl)(CH.sub.2).sub.n-- where n is 0-3 and said cycloalkyl is
optionally substituted with CN, OH, R.sup.aR.sup.bN--, (1-3C)alkyl
or (1-3C)alkoxy, (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3,
(j) hetCyc.sup.2(CH.sub.2).sub.p-- where p is 0 or 1, (k)
hetAr.sup.1(CH.sub.2).sub.q-- where q is 1 or 2, (1) halogen and
(m) hetCyc.sup.1C(.dbd.O)CH.sub.2--;
[0214] hetCyc.sup.1 is a 4-7 membered saturated heterocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterocyclic ring is optionally substituted with one
or more substituents independently selected from the group
consisting of fluoro, HO, C1-C6 alkyl (optionally substituted with
1-3 fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted
with 1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0215] hetCyc.sup.2 is a 7-10 membered heterospirocyclic ring
having 1-2 ring heteroatoms independently selected from N and O,
wherein said heterospirocyclic ring is optionally substituted with
one or more substituents independently selected from the group
consisting of C1-C6 alkyl (optionally substituted with 1-3
fluoros), (C1-C6 alkoxy)C1-C6 alkyl- (optionally substituted with
1-3 fluoros), (C3-C6 cycloalkoxy)C1-C6 alkyl-, hydroxy(C1-C6
alkyl)-, R.sup.cR.sup.dN- and (C1-C6 alkyl)C(.dbd.O)--;
[0216] hetAr.sup.1 is a 6-membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said ring is optionally substituted with
one or more substituents independently selected from C1-C6 alkyl
and halogen;
[0217] R.sup.2 is Ar.sup.1 or hetAr.sup.2;
[0218] Ar.sup.1 is phenyl substituted with one or more groups
independently selected from halogen, cyano, C1-C3 alkyl, C1-C3
alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--, (C1-C3 alkyl)C(.dbd.O)NH--,
(cyclopropyl)C(.dbd.O)NH-- and (cyclopropyl)NHC(.dbd.O)--, wherein
each of said C1-C3 alkyl and C1-C3 alkoxy portions is optionally
substituted with 1-3 fluoros;
[0219] hetAr.sup.2 is a 6-membered heteroaryl ring having 1-2 ring
nitrogen atoms, wherein said heteroaryl ring is optionally
substituted with one or more groups independently selected from
halogen, C1-C3 alkyl, C1-C3 alkoxy, (C1-C3 alkyl)NHC(.dbd.O)--,
(C1-C3 alkyl)C(.dbd.O)NH--, (C3-C4 cycloalkyl)C(.dbd.O)NH-- and
(C3-C4 cycloalkyl)NHC(.dbd.O)--, wherein each of said C1-C3 alkyl
and C1-C3 alkoxy portions is optionally substituted with 1-3
fluoros;
[0220] R.sup.3 is hydrogen, C1-C4 alkyl or (C3-C4)cycloalkyl;
and
[0221] R.sup.a, R.sup.b, R.sup.c and R.sup.d are independently
hydrogen or C1-C6 alkyl optionally substituted with F, OH or C1-C6
alkoxy.
[0222] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is as defined for Formula I-B.
[0223] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkyl (optionally
substituted with 1-3 fluoros), and C1-C3 alkoxy (optionally
substituted with 1-3 fluoros).
[0224] In one embodiment of Formula I-C, R.sup.3 is hydrogen.
[0225] In one embodiment of Formula I-C, R.sup.3 is C1-C4
alkyl.
[0226] In one embodiment of Formula I-C, R.sup.3 is
(C3-C4)cycloalkyl.
[0227] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkoxy
(optionally substituted with 1-3 fluoros) and (C1-C3
alkyl)NHC(.dbd.O)--; and R.sup.3 is hydrogen.
[0228] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkoxy
(optionally substituted with 1-3 fluoros) and (C1-C3
alkyl)NHC(.dbd.O)--; and R.sup.3 is C1-C4 alkyl.
[0229] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkoxy
(optionally substituted with 1-3 fluoros) and (C1-C3
alkyl)NHC(.dbd.O)--; and R.sup.3 is (C3-C4)cycloalkyl.
[0230] In one embodiment of Formula I-C, R.sup.1 is independently
selected from (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros) and (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3; and
hetCyc.sup.1 is defined for Formula I-C.
[0231] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkoxy
(optionally substituted with 1-3 fluoros) and (C1-C3
alkyl)NHC(.dbd.O)--; R.sup.3 is hydrogen; R.sup.1 is independently
selected from (b) C1-C6 alkyl (optionally substituted with 1-3
fluoros) and (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is 0-3; and
hetCyc.sup.1 is defined for Formula I-C.
[0232] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkoxy
(optionally substituted with 1-3 fluoros) and (C1-C3
alkyl)NHC(.dbd.O)--; R.sup.3 is C1-C4 alkyl; R.sup.1 is
independently selected from (b) C1-C6 alkyl (optionally substituted
with 1-3 fluoros) and (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is
0-3; and hetCyc.sup.1 is defined for Formula I-C.
[0233] In one embodiment of Formula I-C, R.sup.2 is Ar.sup.1,
wherein Ar.sup.1 is phenyl optionally substituted with one or more
groups independently selected from halogen, C1-C3 alkoxy
(optionally substituted with 1-3 fluoros) and (C1-C3
alkyl)NHC(.dbd.O)--; R.sup.3 is (C3-C4)cycloalkyl; R.sup.1 is
independently selected from (b) C1-C6 alkyl (optionally substituted
with 1-3 fluoros) and (i) hetCyc.sup.1(CH.sub.2).sub.m-- where m is
0-3; and hetCyc.sup.1 is defined for Formula I-C.
[0234] It will be appreciated that certain compounds provided
herein may contain one or more centers of asymmetry and may
therefore be prepared and isolated in a mixture of isomers such as
a racemic mixture, or in an enantiomerically pure form.
[0235] It will further be appreciated that the compounds of Formula
I or their salts may be isolated in the form of solvates, and
accordingly that any such solvate is included within the scope of
the present invention. For example, compounds of Formula I and
salts thereof can exist in unsolvated as well as solvated forms
with pharmaceutically acceptable solvents such as water, ethanol,
and the like.
[0236] The compounds of Formula I include pharmaceutically
acceptable salts thereof. In addition, the compounds of Formula I
also include other salts of such compounds which are not
necessarily pharmaceutically acceptable salts, and which may be
useful as intermediates for preparing and/or purifying compounds of
Formula I and/or for separating enantiomers of compounds of Formula
I. Non-limiting examples of salts include monochloride, dichloride,
trifluoroacetic acid, and di-trifluoroacetic acid salts of
compounds of Formula I.
[0237] In one embodiment, the compounds of Formula I include the
compounds of Examples 1-62 and stereoisomers and pharmaceutically
acceptable salts and solvates thereof. In one embodiment, the
compounds of Examples 1-83 are in the free base form. In one
embodiment, the compounds of Examples 1-83 are monochloride,
dichloride, trifluoroacetic acid, or di-trifluoroacetic acid
salts.
[0238] The term "pharmaceutically acceptable" indicates that the
substance or composition is compatible chemically and/or
toxicologically, with the other ingredients comprising a
formulation, and/or the patient being treated therewith.
[0239] Compounds provided herein may also contain unnatural
proportions of atomic isotopes at one or more of the atoms that
constitute such compounds. That is, an atom, in particular when
mentioned in relation to a compound according to Formula I,
comprises all isotopes and isotopic mixtures of that atom, either
naturally occurring or synthetically produced, either with natural
abundance or in an isotopically enriched form. For example, when
hydrogen is mentioned, it is understood to refer to .sup.1H,
.sup.2H, .sup.3H or mixtures thereof; when carbon is mentioned, it
is understood to refer to .sup.11C, .sup.12C, .sup.13C .sup.14C or
mixtures thereof; when nitrogen is mentioned, it is understood to
refer to .sup.13N, .sup.14N, .sup.15N or mixtures thereof; when
oxygen is mentioned, it is understood to refer to .sup.14O,
.sup.15O, .sup.16O, .sup.17O, .sup.18O or mixtures thereof; and
when fluoro is mentioned, it is understood to refer to .sup.18F,
.sup.19F or mixtures thereof. The compounds provided herein
therefore also comprise compounds with one or more isotopes of one
or more atom, and mixtures thereof, including radioactive
compounds, wherein one or more non-radioactive atoms has been
replaced by one of its radioactive enriched isotopes. Radiolabeled
compounds are useful as therapeutic agents, e.g., cancer
therapeutic agents, research reagents, e.g., assay reagents, and
diagnostic agents, e.g., in vivo imaging agents. All isotopic
variations of the compounds provided herein, whether radioactive or
not, are intended to be encompassed within the scope of the present
invention.
[0240] For illustrative purposes, Schemes 1 and 1A show general
methods for preparing the compounds provided herein as well as key
intermediates. For a more detailed description of the individual
reaction steps, see the Examples section below. Those skilled in
the art will appreciate that other synthetic routes may be used to
synthesize the inventive compounds. Although specific starting
materials and reagents are depicted in the Schemes and discussed
below, other starting materials and reagents can be easily
substituted to provide a variety of derivatives and/or reaction
conditions. In addition, many of the compounds prepared by the
methods described below can be further modified in light of this
disclosure using conventional chemistry well known to those skilled
in the art.
##STR00026##
[0241] Scheme 1 shows a general scheme for the synthesis of
compound 9 where X, R.sup.1, R.sup.2, R.sup.3, Ring A and z are as
defined for Formula I. Compound 3, where R.sup.3 is as defined for
Formula I, may be obtained by treating compound 1 (commercially
available or prepared according to Scheme 2) with boronic acid 2,
where R.sup.2 is as defined for Formula I, in the presence of
Cu(II) catalyst such as cupric acetate and a ligand such as
pyridine. Compound 3 may be reacted with a dioxoborinane, such as
bis(pinacolato)diboron, using appropriate Suzuki coupling reaction
conditions (e.g., in the presence of a palladium (II) catalyst such
as Pd(OAc).sub.2, Pd.sub.2(dba).sub.3, Pd(PPh.sub.3).sub.4 or
Pd(Dppf).sub.2 and optionally in the presence of a suitable ligand
such as XPhos and in the presence of an inorganic base such as
potassium acetate or sodium carbonate), to provide compound 4 where
R.sup.2 and R.sup.3 are as defined for Formula I. Compound 4 may be
reacted with compound 5 (prepared as described below) where X, Ring
A, R.sup.1 and z are as defined for Formula I, using appropriate
Suzuki coupling reaction conditions (e.g., in the presence of a
palladium (II) catalyst such as Pd(PPh.sub.3).sub.4,
Pd(Dppf).sub.2, Pd(OAc).sub.2, or Pd.sub.2(dba).sub.3, and an
inorganic base such as potassium carbonate or sodium carbonate) to
provide compound 9. Compound 5 may be prepared by reacting compound
6 (where R.sup.1, Ring A and z are as defined for Formula I) with
compound 7 (where X is as defined for Formula I) to provide
compound 8, which may subsequently be brominated using standard
conditions to provide compound 5. The syntheses of intermediates 3
and 6 which are not commercially available are described in the
Examples.
##STR00027##
[0242] Scheme 1A shows a general scheme for the synthesis of
compound 9B where X, R.sup.2 and R.sup.3 are as defined for Formula
I, z is 1 and R.sup.1 is a piperidine substituted with C1-C6 alkyl
(optionally substituted with 1-3 fluoros), (C1-C3 alkoxy)C1-C6
alkyl (optionally substituted with 1-3 fluoros) or hydroxy(C1-C6
alkyl). Compound 9A, where X, R.sup.2 and R.sup.3 are as defined
for Formula I, z is 1 and R.sup.1 is a piperidine substituted with
C1-C6 alkyl (optionally substituted with 1-3 fluoros), (C1-C3
alkoxy)C1-C6 alkyl (optionally substituted with 1-3 fluoros) or
hydroxy(C1-C6 alkyl), prepared as described in Scheme 1, may be
reacted with a compound having the formula R.sup.x--Y, where
R.sup.x is C1-C6 alkyl (optionally substituted with 1-3 fluoros),
(C1-C3 alkoxy)C1-C6 alkyl (optionally substituted with 1-3 fluoros)
or hydroxy(C1-C6 alkyl), and Y is a leaving group such as a halogen
or a tosylate, under standard alkylation reaction conditions, for
example in the presence of an inorganic base such as potassium
carbonate, to provide compound 9B.
[0243] The compound of formulas 1, 3, 4, 5, 6, 8 and 9A as shown
and described above for Schemes 1 and 1A are useful as
intermediates for preparing compounds of Formula I and are provided
as further aspects of the invention.
[0244] Further provided herein is a process for preparing of a
compound of Formula I or a pharmaceutically acceptable salt thereof
as defined herein which comprises:
[0245] (a) reacting a compound having the formula 5:
##STR00028##
[0246] where X, Ring A, R.sup.1 and z are as defined for Formula I,
with a compound having the formula 4:
##STR00029##
[0247] where R.sup.2 is as defined for Formula I, in the presence
of a palladium (II) catalyst and an inorganic base; or
[0248] (b) for a compound of Formula I where X, R.sup.2 and R.sup.3
are as defined for Formula I, z is 1 and R.sup.1 is a piperidine
substituted with C1-C6 alkyl (optionally substituted with 1-3
fluoros), (C1-C3 alkoxy)C1-C6 alkyl (optionally substituted with
1-3 fluoros) or hydroxy(C1-C6 alkyl), reacting a compound having
the formula 9A:
##STR00030##
[0249] where X, R.sup.2 and Ring A are as defined for Formula I,
with a compound having the formula R.sup.x--Y, where R.sup.x is
C1-C6 alkyl (optionally substituted with 1-3 fluoros), (C1-C3
alkoxy)C1-C6 alkyl (optionally substituted with 1-3 fluoros) or
hydroxy(C1-C6 alkyl), and Y is a leaving group such as a halogen or
a tosylate, under standard alkylation reaction conditions; and
[0250] removing any protecting groups if present and optionally
forming a pharmaceutically acceptable salt.
[0251] The ability of test compounds to act as inhibitors of FGFR1,
FGFR2 and/or FGFR3 may be demonstrated by the assay described in
Example A. IC.sub.50s are shown in Table F.
[0252] Compounds of Formula I have been found to inhibit FGFR1,
FGFR2 and/or FGFR3, and are therefore believed to be useful for
treating diseases and disorders which can be treated with an
inhibitor of FGFR1, FGFR2, FGFR3 and/or FGFR4, such as
FGFR-associated diseases and disorders, e.g., proliferative
disorders such as cancers, including hematological cancers and
solid tumors.
[0253] In certain embodiments, compounds of Formula I are useful
for preventing diseases and disorders as defined herein (for
example cancer).
[0254] The term "preventing" as used herein means the prevention of
the recurrence or spread, in whole or in part, of the disease or
condition as described herein, or a symptom thereof.
[0255] As used herein, the word "a" before a noun represents one or
more of the particular noun. For example, the phrase "a cell"
represents "one or more cells."
[0256] As used herein, terms "treat" or "treatment" refer to
therapeutic or palliative measures. Beneficial or desired clinical
results include, but are not limited to, alleviation, in whole or
in part, of symptoms associated with a disease or disorder or
condition, diminishment of the extent of disease, stabilized (i.e.,
not worsening) state of disease, delay or slowing of disease
progression, amelioration or palliation of the disease state (e.g.,
one or more symptoms of the disease), and remission (whether
partial or total). "Treatment" can also mean prolonging survival as
compared to expected survival if not receiving treatment.
[0257] The term "FGFR-associated disease or disorder" as used
herein refers to diseases or disorders associated with or having a
dysregulation of a FGFR gene, a FGFR protein, or the expression or
activity, or level of the same (e.g., one or more of the same)
(e.g., any of the types of dysregulation of an FGFR gene, a FGFR
protein, or expression or activity, or level of the same, described
herein). A non-limiting example of an FGFR-associated disease or
disorder is an FGFR-associated cancer.
[0258] As used herein, the term "FGFR-associated cancer" shall be
defined to include cancers associated with or having dysregulation
of a FGFR gene, a FGFR protein, or expression or activity, or level
of the same (e.g., any of types of dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same,
described herein). Non-limiting examples of a FGFR-associated
cancer are described herein.
[0259] As used herein, the term "subject," "individual," or
"patient," used interchangeably, refers to any animal, including
mammals such as mice, rats, other rodents, rabbits, dogs, cats,
swine, cattle, sheep, horses, primates, and humans. In some
embodiments, the patient is a human. In some embodiments, the
subject has experienced and/or exhibited at least one symptom of
the disease or disorder to be treated and/or prevented. In some
embodiments, the subject has been identified or diagnosed as having
a cancer with dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same (a FGFR-associated
cancer) (e.g., as determined using a regulatory agency-approved,
e.g., FDA-approved, assay or kit). In some embodiments, the subject
has a tumor that is positive for dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same
(e.g., as determined using a regulatory agency-approved assay or
kit). The subject can be a subject with a tumor(s) that is positive
for dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same (e.g., identified as positive using
a regulatory agency-approved, e.g., FDA-approved, assay or kit).
The subject can be a subject whose tumors have dysregulation of a
FGFR gene, a FGFR protein, or expression or activity, or a level of
the same (e.g., where the tumor is identified as such using a
regulatory agency-approved, e.g., FDA-approved, kit or assay). In
some embodiments, the subject is suspected of having a
FGFR-associated cancer. In some embodiments, the subject has a
clinical record indicating that the subject has a tumor that has
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same (and optionally the clinical record
indicates that the subject should be treated with any of the
compositions provided herein).
[0260] The term "FGFR" or "FGFR protein" includes any of the FGFR
proteins described herein (e.g., a FGFR1, a FGFR2, a FGFR3 or a
FGFR4 protein, or isoforms thereof).
[0261] The term "FGFR gene" includes any of the FGFR genes
described herein (e.g., a FGFR1, a FGFR2, a FGFR3 gene, or a FGFR4
gene).
[0262] The term "wildtype" or "wild-type" describes a nucleic acid
(e.g., a FGFR gene or a FGFR mRNA) or protein (e.g., a FGFR
protein) that is found in a subject that does not have a
FGFR-associated disease, e.g., a FGFR-associated cancer (and
optionally also does not have an increased risk of developing a
FGFR-associated disease and/or is not suspected of having a
FGFR-associated disease), or is found in a cell or tissue from a
subject that does not have a FGFR-associated disease, e.g., a
FGFR-associated cancer (and optionally also does not have an
increased risk of developing a FGFR-associated disease and/or is
not suspected of having a FGFR-associated disease).
[0263] The term "regulatory agency" is a country's agency for the
approval of the medical use of pharmaceutical agents with the
country. For example, a non-limiting example of a regulatory agency
is the U.S. Food and Drug Administration (FDA).
[0264] The phrase "dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same" is a genetic mutation
(e.g., a FGFR gene translocation that results in the expression of
a fusion protein, a deletion in a FGFR gene that results in the
expression of a FGFR protein that includes a deletion of at least
one amino acid as compared to the wild-type FGFR protein, or a
mutation in a FGFR gene that results in the expression of a FGFR
protein with one or more point mutations, an alternative spliced
version of a FGFR mRNA that results in a FGFR protein that results
in the deletion of at least one amino acid in the FGFR protein as
compared to the wild-type FGFR protein), or a FGFR gene
amplification that results in overexpression of a FGFR protein) or
an autocrine activity resulting from the overexpression of a FGFR
gene a cell, that results in a pathogenic increase in the activity
of a kinase domain of a FGFR protein (e.g., a constitutively active
kinase domain of a FGFR protein) in a cell. For example, a
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, can be a mutation in a FGFR1,
FGFR2, FGFR3, or FGFR4 gene that encodes a FGFR protein that is
constitutively active or has increased activity as compared to a
protein encoded by a FGFR1, FGFR2, FGFR3, or FGFR4 gene that does
not include the mutation. For example, a dysregulation of a FGFR
gene, a FGFR protein, or expression or activity, or level of the
same, can be the result of a gene or chromosome translocation which
results in the expression of a fusion protein that contains a first
portion of FGFR1, FGFR2, FGFR3, or FGFR4 that includes a functional
kinase domain, and a second portion of a partner protein (i.e.,
that is not FGFR1, FGFR2, FGFR3, or FGFR4). In some examples,
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, can be a result of a gene translation of one FGFR1 gene
with another FGFR1 gene. Non-limiting examples of fusion proteins
that are a result of a FGFR gene translocation are described in
Table 3.
[0265] A dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same, can, e.g., include a
mutation(s) in a FGFR1, FGFR2, FGFR3, or FGFR4 gene that results in
a FGFR1, FGFR2, FGFR3, or FGFR4 protein containing at least one
(e.g., two, three, four, or five) point mutations (e.g., one of
more of the point mutations listed in Table 1).
[0266] A dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same, can be a mutation in
a FGFR1, FGFR2, FGFR3, or FGFR4 gene that results in a deletion of
one or more contiguous amino acids (e.g., at least two, at least
three, at least four, at least 5, at least 6, at least 7, at least
8, at least 9, at least 10, at least 15, at least 20, at least 30,
at least 40, at least 50, at least 60, at least 70, at least 80, at
least 90, at least 100, at least 110, at least 120, at least 130,
at least 140, at least 150, at least 160, at least 170, at least
180, at least 190, at least 200, at least 210, at least 220, at
least 230, at least 240, at least 250, at least 260, at least 270,
at least 280, at least 290, at least 300, at least 310, at least
320, at least 330, at least 340, at least 350, at least 360, at
least 370, at least 380, at least 390, or at least 400 amino acids)
in the FGFR1, FGFR2, FGFR3, or FGFR4 protein (except for the
deletion of amino acids in the kinase domain of FGFR1, FGFR2,
FGFR3, or FGFR4 that would result in inactivation of the kinase
domain).
[0267] In some examples, a dysregulation of a FGFR gene, a FGFR
protein, or expression or activity, or level of the same, can
include an alternate spliced form of a FGFR mRNA. In some examples,
a dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, includes an amplification of a FGFR
gene (e.g., one, two, three, or four additional copies of a FGFR1,
FGFR2, FGFR3, and/or FGFR4 gene) that can result, e.g., in an
autocrine expression of a FGFR gene in a cell.
[0268] The term "mammal" as used herein, refers to a warm-blooded
animal that has or is at risk of developing a disease described
herein and includes, but is not limited to, guinea pigs, dogs,
cats, rats, mice, hamsters, and primates, including humans.
[0269] The phrase "time of survival" means the length of time
between the identification or diagnosis of cancer (e.g., any of the
cancers described herein) in a subject or patient by a medical
professional and the time of death of the subject or patient
(caused by the cancer). Methods of increasing the time of survival
in a subject or patient having a cancer are described herein.
[0270] The term "metastasis" is an art known term and means the
formation of an additional tumor (e.g., a solid tumor) at a site
distant from a primary tumor in a subject or patient, where the
additional tumor includes the same or similar cancer cells as the
primary tumor.
[0271] The phrase "risk of developing a metastasis" means the risk
that a subject or patient having a primary tumor will develop an
additional tumor (e.g., a solid tumor) at a site distant from a
primary tumor in a subject or patient over a set period of time,
where the additional tumor includes the same or similar cancer
cells as the primary tumor. Methods for reducing the risk of
developing a metastasis in a subject or patient having a cancer are
described herein.
[0272] The phrase "risk of developing additional metastases" means
the risk that a subject or patient having a primary tumor and one
or more additional tumors at sites distant from the primary tumor
(where the one or more additional tumors include the same or
similar cancer cells as the primary tumor) will develop one or more
further tumors distant from the primary tumor, where the further
tumors include the same or similar cancer cells as the primary
tumor. Methods for reducing the risk of developing additional
metastasis are described herein.
[0273] The term "angiogenesis-related disorder" means a disease
characterized in part by an increased number or size of blood
vessels in a tissue in a subject or patient, as compared to a
similar tissue from a subject not having the disease. Non-limiting
examples of angiogenesis-related disorders include: cancer (e.g.,
any of the exemplary cancers described herein, such as prostate
cancer, lung cancer, breast cancer, bladder cancer, renal cancer,
colon cancer, gastric cancer, pancreatic cancer, ovarian cancer,
melanoma, hepatoma, sarcoma, and lymphoma), exudative macular
degeneration, proliferative diabetic diabetic retinopathy, ischemic
retinopathy, retinopathy of prematurity, neovascular glaucoma,
iritis rubeosis, corneal neovascularization, cyclitis, sickle cell
retinopathy, and pterygium.
[0274] The term "resistant cancer cell to an anti-cancer drug"
means a cancer cell that demonstrates an increased rate of growth
and/or proliferation in the presence of an anti-cancer drug as
compared to the rate of growth and/or proliferation of a similar
cancer cell (or an average rate of growth and/or proliferation of a
population of a similar cancer cells). For example, a cancer cell
that demonstrates an increased rate of growth and/or proliferation
in the presence of an anti-cancer drug (as compared to the rate of
growth and/or proliferation of a similar cancer cell) can be
present in a patient or a subject (e.g., a patient or a subject
having a FGFR-associated cancer).
[0275] The term "increasing sensitivity to an anti-cancer drug"
means a decrease in the rate of growth and/or proliferation of a
resistant cancer cell (to an anti-cancer drug) when contacted with
the anti-cancer drug and at least one of the compounds described
herein, as compared to the rate of growth and/or proliferation of a
resistant cancer cell when contacted with the anti-cancer drug
alone.
[0276] The FGFR receptors (FGFR1, FGFR2, FGFR3, and FGFR4) share
several structural features in common, including three
extracellular immunoglobulin-like (Ig) domains, a hydrophobic
transmembrane domain, and an intracellular split tyrosine kinase
domain with a 14-amino acid insertion (Johnson et al., Adv. Cancer
Res. 60:1-40, 1993; and Wilkie et al., Curr. Biol. 5:500-507,
1995). Several isoforms of each FGFR have been identified and are
the result of alternative splicing of their mRNAs (Johnson et al.,
Mol. Cell. Biol. 11:4627-4634, 1995; and Chellaiah et al., J. Biol.
Chem. 269:11620-11627, 1994). A few of the receptor variants that
result from this alternative splicing have different ligand binding
specificities and affinities (Zimmer et al., J. Biol. Chem.
268:7899-7903, 1993; Cheon et al., Proc. Natl. Acad. Sci. U.S.A.
91:989-993, 1994; and Miki et al., Proc. Natl. Acad. Sci. U.S.A.
89:246-250, 1992). Protein sequences for FGFR proteins and nucleic
acids encoding FGFR proteins are known in the art. Exemplary amino
acid sequences for exemplary wildtype isoforms of FGFR1 are SEQ ID
NO: 1 and SEQ ID NO: 2. Exemplary amino acid sequences for
exemplary wildtype isoforms of FGFR2 are SEQ ID NO: 2 and SEQ ID
NO: 3. Exemplary amino acid sequences for exemplary wildtype
isoforms of FGFR3 are SEQ ID NO: 5 and SEQ ID NO: 6. Exemplary
amino acid sequences for exemplary wildypte isoforms of FGFR4 are
SEQ ID NO: 7 and SEQ ID NO: 8.
[0277] Signaling by FGFRs regulates key biological processes
including cell proliferation, survival, migration, and
differentiation. Dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same, has been associated
with many types of cancer. For example, dysregulation of FGFRs can
occur by multiple mechanisms, such as FGFR gene overexpression,
FGFR gene amplification, activating mutations (e.g., point
mutations or truncations), and chromosomal rearrangements that lead
to FGFR fusion proteins. Dysregulation of a FGFR gene, a FGFR
protein, or expression or activity, or level of the same, can
result in (or cause in part) the development of a variety of
different FGFR-associated cancers. Non-limiting examples of the
types of FGFR-associated cancers and the dysregulation of a FGFR
gene, a FGFR protein, or expression or activity, or level of the
same, that causes (or causes in part) the development of the
FGFR-associated cancers are listed in Tables A-D.
[0278] For example, dysregulation of a FGFR1 gene, a FGFR1 protein,
or expression or activity, or level of the same, can include FGFR1
gene amplification, a FGFR1 gene fusion from those listed in Table
C, and/or one or more point mutations selected from those listed in
Table A (e.g., one of more of T141R, R445W, N546K, K656E, and
G818R). Dysregulation of a FGFR2 gene, a FGFR2 protein, or
expression or activity, or level of the same, can, e.g., include
FGFR2 gene amplification, a FGFR2 gene fusion from those listed in
Table C, and/or one or more point mutations selected from those
listed in Table A (e.g., one or more of S252W, P253R, A315T, D336N,
Y375C, C382R, V395D, D471N, I547V, N549K, N549K, N549Y, and
K659E).
[0279] Dysregulation of a FGFR3 gene, a FGFR3 protein, or
expression or activity, or level of the same can, e.g., include
FGFR3 gene amplification, a FGFR3 gene fusion from those listed in
Table C, and/or one or more point mutations selected from those
listed in Table A (e.g., one or more of S131L, R248C, S249C, G370C,
S371C, Y373C, G380R, R399C, E627K, K650E, K650M, V677I, and D785Y).
Dysregulation of a FGFR4 gene, a FGFR4 protein, or expression or
activity, or level of the same can, e.g., include FGFR4 gene
amplification and/or one or more point mutations selected from
those listed in Table A (e.g., one or more of R183S, R394Q, D425N,
V510L, and R610H).
[0280] Additional examples of FGFR fusion proteins, FGFR point
mutations, FGFR gene overexpression, or FGFR gene amplification
that cause (or cause in part) the development of a FGFR-associated
cancer are described in: Wu et al., Cancer Discovery 3:636, 2013;
Wesche et al., Biochem. J. 437:199-213, 2011; Gallo et al.,
Cytokine Growth Factor Rev. 26:425-449, 2015; Parker et al., J.
Pathol. 232:4-15, 2014; Katoh et al., Expert Rev. Anticancer Res.
10:1375-1379, 2010; Chang et al., PLoS One 9:e105524, 2014;
Kelleher et al., Carcinogenesis 34:2198-2205, 2013; Katoh et al.,
Med. Res. Rev. 34:280-300, 2014; Knights et al., Pharmacol.
Therapeutics 125:105-117, 2010; Turner et al., Sci. Transl. Med.
2:62ps56, 2010; Dutt et al., PLoS One 6(6):e20351, 2011; Weiss et
al., Sci. Transl. Med. 2:62ra93, 2010; Becker et al., J.
Neurophatol. Exp. Neurol. 74:743-754, 2015; Byron et al., PLoS One
7(2):e30801, 2012; van Rhihn et al., Eur. J. Human Genetics
10:819-824, 2002; Hart et al., Oncogene 19(29):3309-3320, 2000; Lin
et al., Cancer Res. 68:664-673, 2008; and Helsten et al., Clin.
Cancer Res., e-publication dated Sep. 15, 2015 (each of which is
incorporated herein by reference). Additional non-limiting aspects
and examples of FGFR fusion proteins, FGFR point mutations, FGFR
gene overexpression, or FGFR gene amplification are described
below.
[0281] Point Mutations
[0282] FGFR mutations that confer constitutive activation have been
described in a number of congenital skeletal disorders (Turner N,
Grose R., Nat Rev Cancer 2010; 10:116-129). FGFRs have been
identified as among the most commonly mutated kinase genes in human
cancers, with mutations in FGFR2 and FGFR3 being most prevalent
(Turner N., Grose R., Nat Rev Cancer 2010; 10:116-129). For
example, approximately 50% to 60% of non-muscle invasive and 17% of
high-grade bladder cancers possess FGFR3 mutations that cause
constitutive FGFR dimerization and activation (Cappellen D. et al.,
Nat Genet 1999; 23:18-20). Activating and oncogenic FGFR2 mutations
located in the extracellular and kinase domains of the receptor
have been described in 12% of endometrial carcinomas (Dutt A. et
al., Proc Natl Acad Sci USA 2008; 105:8713-8717). Importantly, the
FGFR2 mutations found in endometrial cancer confer sensitivity to
FGFR inhibition (Dutt A. et al., Proc Natl Acad Sci USA 2008;
105:8713-8717). More recently, FGFR2 mutations have been described
in 5% of squamous non-small cell lung cancers (NSCLC; Hammerman P.
et al., Genomic characterization and targeted therapeutics in
squamous cell lung cancer [abstract]. In: Proceedings of the 14th
World Conference on Lung Cancer; 2011 3-7 July; Aurora (Colo.):
International Association for the Study of Lung Cancer; 2011).
FGFR3 mutations in bladder cancer and FGFR2 mutations in
endometrial cancer are mutually exclusive with mutations in HRAS
and KRAS, respectively. In addition, mutations in the FGFR4 kinase
domain have been found in the childhood soft tissue sarcoma
rhabdomyosarcoma, causing autophosphorylation and constitutive
signaling (Taylor J G, et al., J Clin Invest 2009; 119:3395-407).
FGFR1, FGFR2, FGFR3, and/or FGFR4 can include one, two, three,
four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen,
fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or
twenty different point mutations (as compared to an appropriate
corresponding wildtype FGFR1, FGFR2, FGFR3, or FGFR4 amino acid
sequence, respectively). Non-limiting examples of point mutations
in FGFR1, FGFR2, FGFR3, or FGFR4 that are thought to cause (or
cause in-part) a FGFR-associated cancer are listed in Table A.
FGFR Gene Amplification
[0283] FGFR gene amplification often leads to FGFR overexpression,
which can provoke ligand-independent signaling. In breast cancer,
amplification of the genomic locus of FGFR1 (8p11-12) occurs in
approximately 10% of predominantly estrogen receptor (ER)-positive
patients (Taylor J G, et al., J Clin Invest 2009; 119:3395-4307).
In vitro studies support the potential oncogenic nature of FGFR1
amplification (Welm B E, et al., J Cell Biol 2002; 157:703-14);
however, due to the gene-dense nature of the 8p11-12 amplicon in
breast cancer, there is continuing debate about the identity of the
driving oncogene. More recently. FGFR1 has been found to be
amplified in 22% of squamous NSCLC (Weiss J, et al., Sci Transl Med
2010; 2:62ra93), and these amplifications seem to confer dependence
upon FGFR signaling. Unlike the broad amplicon containing FGFR1
found in breast cancers, the amplicon in lung is more focal; it
remains to be seen if these differences influence the degree of
oncogenic addiction to FGFR1. FGFR2 amplifications have been
reported in up to 10% of gastric cancers, most of which are
diffuse-type with relatively poor prognosis (Kunii K, et al.,
Cancer Res 2008; 68:2340-2348). Further, in an FGFR2-amplified
gastric cancer cell line, Snu-16, FGFR2 downregulation led to
significant inhibition of cell growth and survival that further
translated into tumor growth regression in vivo (Xie L, et al.,
AZD4547, a potent and selective inhibitor of FGF-receptor tyrosine
kinases 1, 2 and 3, inhibits the growth of FGF-receptor 2 driven
gastric cancer models in vitro and in vivo. In: Proceedings of the
American Association of Cancer Research Annual Meeting; 2011 Apr.
2-6; Orlando (Fla.). Philadelphia (Pa.): AACR; 2011. Abstract nr
1643). In some gastric cancer cell lines, FGFR2 amplification is
accompanied by deletion of the coding exon located proximal to the
C-terminus (Ueda T, et al., Cancer Res 1999; 59:6080-6086). This
deletion impedes receptor internalization, thereby contributing to
constitutive activation of the receptor. The presence of FGFR2 gene
amplifications in gastric cancer is associated with sensitivity to
inhibition of FGFR signaling by tyrosine kinase inhibitors and
monoclonal antibodies in preclinical models (Zhao G, et al., Mol
Cancer Ther 2011; 10:2200-2210; Zhao W M, et al., Clin Cancer Res
2010; 16:5750-5758). Non-limiting examples of FGFR-associated
cancers that are caused (or caused in-part) by the amplification
and/or overexpression of the FGFR1 gene, the FGFR2 gene, the FGFR3
gene, or the FGFR4 gene are listed in Table B.
TABLE-US-00001 TABLE B Overexpression or Amplification of FGFR
Genes and FGFR-Associated Cancer FGFR Gene Type of Dysregulation
FGFR-Associated Cancer FGFR1 Amplification or Breast cancer or
carcinoma (e.g., Overexpression hormone receptor-positive breast
cancer, ductal carcinoma in situ (breast)), pancreatic ductal
adenocarcinoma, pancreatic exocrine carcinoma, smoking- associated
lung cancer, small cell lung cancer, lung adenocarcinoma, non-small
cell lung cancer, squamous cell lung cancer or carcinoma, prostate
cancer or carcinoma, ovarian cancer, fallopian tube carcinoma,
bladder cancer, rhabdomyosarcoma, head and neck carcinoma (e.g.,
head and neck squamous cell carcinoma), esophageal cancer (e.g.,
esophageal squamous cell carcinoma), sarcoma (e.g., osteosarcoma),
hepatocellular carcinoma, renal cell carcinoma, colorectal cancer
(e.g., colorectal adenocarcinoma), prostate cancer, salivary gland
tumors, glioblastoma multiforme, urinary bladder cancer, urothelial
carcinoma, carcinoma of unknown primary, squamous non- lung tumors,
gastric cancer, gastroesophageal junction carcinoma, adenoid cystic
carcinoma, anal squamous cell carcinoma, oral squamous cell
carcinoma, cholangiocarcinoma, hemangioendothelioma,
leiomyosarcoma, melanoma, neuroendocrine carcinoma, squamous cell
carcinoma, uterine carcinosarcoma FGFR2 Amplification Gastric
cancer, gastroesophageal junction adenocarcinoma, breast cancer
(e.g., triple-negative breast cancer), colon cancer, colorectal
cancer (e.g., colorectal adenocarcinoma), urothelial cancer,
bladder adenocarcinoma, carcinoma of unknown primary,
cholangiocarcinoma, endometrial adenocarcinoma, esophageal
adenocarcinoma, gallbladder carcinoma, ovarian cancer, fallopian
tube carcinoma, pancreatic exocrine carcinoma, sarcoma, squamous
cell carcinoma FGFR2 Overexpression Myxoid lipocarcinoma, rectal
cancer, renal cell carcinoma, breast cancer FGFR3 Upregulation of
Activity Colorectal cancer, hepatocellular carcinoma, pancreatic
exocrine carcinoma FGFR3 Overexpression Multiple myeloma, thyroid
carcinoma, FGFR3 Amplification Bladder cancer and salivary adenoid
cystic cancer, urothelial cancer, breast cancer, carcinoid,
carcinoma of unknown primary, colorectal cancer (e.g., colorectal
adenocarcinoma), gallbladder carcinoma, gastric cancer,
gastroesophageal junction adenocarcinoma, glioma, mesothelioma,
non-small cell lung carcinoma, small cell lung cancer, ovarian
cancer, fallopian tube carcinoma, pancreatic exocrine carcinoma
FGFR4 Amplification Rhabdomyosarcoma, prostate cancer or carcinoma,
breast cancer, urothelial cancer, carcinoid, carcinoma of unknown
primary, esophageal adenocarcinoma, head and neck carcinoma,
hepatocellular carcinoma, non-small cell lung carcinoma, ovarian
cancer, fallopian tube carcinoma, peritoneal carcinoma, renal cell
carcinoma FGFR4 Upregulation of Activity Colorectal cancer,
hepatocellular carcinoma, adrenal carcinoma, breast cancer FGFR4
Overexpression Pancreatic intraepithelial neoplasia, and pancreatic
ductal adenocarcinoma
[0284] Fusion Proteins
[0285] Several FGFR translocations have been identified to play a
role in defects in development and in a wide range of malignancies,
whereby chromosomal rearrangement results in a nucleic acid
sequence encoding a fusion protein that includes a kinase domain of
an FGFR protein and an amino acid sequence from a partner protein.
In some examples, fusion proteins are located in the cytosol, do
not undergo lysosomal degradation, are not susceptible to feedback
inhibition, and are permanently dimerized in the absence of ligand.
Such translocations can lead to FGFR overexpression, permanent
dimerization of the fusion protein-FGFR complex, and continuous
signaling. The mechanism of proliferation is dependent on the type
of fusion protein and seems to be disease specific (Jackson C C, et
al., Hum Pathol 2010; 41:461-476). For example, a t(4;14)
intergenic translocation, bringing FGFR3 and the adjacent Multiple
Myeloma SET domain (MMSET) gene under the control of the Ig heavy
chain (IGH) promoter, has been identified in 10% to 20% of multiple
myelomas and is associated with poor prognosis and dependence upon
FGFR signaling (Chesi M, et al., Nat Genet 1997; 16:260-264; Qing
J, et al., J Clin Invest 2009; 119:1216-1229). FGFR3 translocations
are rarely found in prodromal conditions of multiple myeloma,
implicating these translocations in the conversion to full multiple
myeloma. Additional examples of FGFR fusion proteins and the
specific FGFR-associated cancers that they cause (or cause in part)
are listed in Table C. The expression of FGFR fusion proteins can,
e.g., cause (or cause in part) cholangiocarcinoma, bladder cancer,
lung cancer, and breast cancer. Additional examples of FGFR fusion
proteins are known in the art.
[0286] Autocrine and Paracrine Signaling
[0287] Although many of the mechanisms discussed so far are the
result of genetic dysregulation of the FGF/FGFR signaling axis,
ligand-dependent signaling is also likely to play a key role in
cancer development (e.g., described as "Upregulation of Activity"
in Table 2). Autocrine FGF overproduction has been reported in many
tumor types (Turner N, Grose R., Nat Rev Cancer 2010; 10:116-129).
In vitro studies have shown that FGF5 overexpression has been
associated with a number of tumor cell lines (lung, esophagus,
melanoma, colon, and prostate; Hanada K, et al., Cancer Res 2001;
61:5511-5516), and in hepatocellular carcinomas (HCC), the
upregulation of FGF2, 8, 17, and 18 initiates autocrine growth
stimulation, cell survival, and neoangiogenesis (Uematsu S, et al.,
J Gastroenterol Hepatol 2005; 20:583-588; Hu M C, et al., Mol Cell
Biol 1998; 18:6063-6074; Kin M, et al., J Hepatol 1997; 27:677-687;
Gauglhofer C, et al., Hepatology 2011; 53:854-864). Further, HCC
has been found to develop in transgenic mice overexpressing the
hormonal FGF19 (Nicholes K, et al., Am J Pathol 2002;
160:2295-2307), and FGF19 is found on an amplicon on chromosome 11q
that also invariably contains the adjacent FGF3, FGF4, and Cyclin
D1 (CCND1) genes. This amplicon is found in various diseases,
including head and neck squamous cell carcinoma, breast cancer, and
squamous NSCLC. Although there is uncertainty about the key
oncogenic gene on this amplicon or a presumption that it is CCND1,
genetic knockdown of FGF19 inhibits the growth of HCC cell lines
carrying the amplicon (Sawey E T, et al., Cancer Cell 2011;
19:347-358). Autocrine FGF2-FGFR1 feedback loops have also been
reported in NSCLC cell lines and in human melanomas grown as
subcutaneous tumors in nude mice (Marek L, et al., Mol Pharmacol
2009; 75:196-207; Wang Y, Becker D., Nat Med 1997; 3:887-893).
[0288] Paracrine production of FGFs has also been reported in
multiple tumor types. High levels of serum FGF2 have been observed
in small cell lung cancer and are associated with a poor prognosis
(Ruotsalainen T, et al., Cancer Epidemiol Biomarkers Prev 2002;
11:1492-1495), possibly because of an FGF2-mediated cytoprotective
effect, whereby the expression of antiapoptotic proteins are
upregulated, promoting resistance to current anticancer treatments
(Pardo O E, et al., EMBO J 2006; 25:3078-3088). Increased paracrine
expression of one or more of FGF1, 2, 4, 5, 8, and 18 has been
found to promote tumor neoangiogenesis in preclinical models via
the main endothelial FGFRs, FGFR1 and 2 (Presta M, et al., Cytokine
Growth Factor Rev 2005; 16:159-178). Poor prognosis has been
associated with neoangiogenesis in ovarian cancer and melanomas
(Birrer M J, et al., J Clin Oncol 2007; 25:2281-2287).
[0289] Altered FGFR mRNA Splicing
[0290] In addition to overexpression of FGFs, altered splicing of
FGFR mRNAs is another mechanism by which ligand-dependent signaling
is upregulated. Altered FGFR mRNA splicing can allow tumor cells to
be stimulated by a broader range of FGFs than would be capable
under normal physiologic conditions (Zhang X, et al., J Biol Chem
2006; 281:15694-15700). Altered splicing of the IgIII domains in
FGFRs 1, 2, and 3 can switch receptor binding affinity in cancer
cells towards FGFs found in the healthy stroma, creating an
aberrant paracrine signaling loop (Wesche J, Haglund K, Haugsten E
M. et al., Biochem J 2011; 437:199-213). In bladder and prostate
cancer cell lines, a switch from the FGFR2-IIIb isoform to the IIIc
isoform has been associated with tumor progression,
epithelial-mesenchymal transition, and increased invasiveness
(Wesche J, et al., Biochem J 2011; 437:199-213).
[0291] Non-limiting examples of FGFR-associated cancers include
urothelial carcinoma, breast carcinoma or cancer (e.g., hormone
receptor-positive breast cancer, triple-negative breast cancer,
neuroendocrine carcinoma of the breast, mammary carcinoma),
endometriod endometrial cancer or endometrial cancer (e.g.,
endometrial adenocarcinoma), ovarian carcinoma or cancer (e.g.,
ovarian serous cancer), brain cancer (e.g., glioneural tumors,
glioma, pilocytic astrocytoma, rosette-forming glioneural tumor),
cholangiocarcinoma (e.g., intrahepatic cholangiocarcinoma,
metastatic cholangiocarcinoma, medulloblastoma), gastric or stomach
cancer (e.g., gastric adenocarcinoma), gastrointestinal stromal
tumors, lung cancer (e.g., non-small cell lung carcinoma or lung
large cell carcinoma, smoking-associated lung cancer, small cell
lung cancer, lung adenocarcinoma, squamous cell lung cancer or
carcinoma, lung neuroendocrine carcinoma), pancreatic cancer (e.g.,
pancreatic exocrine carcinoma, pancreatic ductal adenocarcinoma,
pancreatic intraepithelial neoplasia), prostate cancer, colorectal
carcinoma or cancer, rectal cancer, renal cell carcinoma,
neuroendocrine carcinoma, head and neck (squamous) carcinoma or
head and neck adenoid cystic carcinoma, skin cancer (e.g.,
melanoma), leiomyosarcoma, sarcoma (e.g., osteosarcoma or soft
tissue sarcoma), osteosarcoma, bladder cancer, uterine cancer,
urinary bladder cancer, rhabdomyosarcoma (e.g., alveolar
rhabdomyosarcoma or embryonal rhabdomyosarcoma), esophageal cancer
(e.g., esophageal adenocarcinoma), hepatocellular carcinoma or
liver cancer, biliary tract cancer, salivary gland tumors (e.g.,
pleomorphic salivary gland adenocarcinoma), glioblatoma multiforme,
myxoid lipocarcinoma, oral cancer (e.g., oral squamous cell
carcinoma), thyroid cancer or carcinoma, anaplastic thyroid
carcinoma, adenoid cystic carcinoma (e.g., salivary adenoid cystic
cancer), glioblastoma multiforme, myeloproliferative
disorders/hematological malignancies (e.g., 8p11 myeloproliferative
syndrome, lymphoma (e.g., T-lymphoblastic lymphoma, T-cell
lymphoma, B-cell lymphoma), leukemia (e.g., acute lymphoblatic
leukemia (ALL), chronic myelogenous leukemia (CML), acute myeloid
leukemia (AML)), myeloproliferative neoplasm, myeloid and lymphoid
neoplasms, stem cell myeloproliferative disorders,
myeloproliferative disorder stem cell leukemiallymphoma syndrome,
chronic myeloid disorder, myeloma (e.g., multiple myeloma)),
phosphaturic mesenchymal tumor, cervical cancer (e.g., cervical
squamous cell carcinoma), gallbladder cancer, spermatocytic
seminoma, seborrheic keratosis, testicular cancer, mesothelioma,
dysembryoplastic neuroepithelial tumor, and dedifferentiated
liposarcoma. Additional examples of FGFR-associated cancers are
listed in Tables 1-3.
[0292] Non-limiting examples of additional FGFR-associated diseases
that are caused by dysregulation of FGFR are listed in Table D. A
subject having any of the additional FGFR-associated diseases
described herein or known in the art can be treated by
administering to the subject a therapeutically effective amount of
a compound of General Formula I (e.g., any of the exemplary
compounds described herein).
[0293] Additional point mutations in FGFR1, FGFR2, FGFR3, and FGFR4
have been identified to result in resistance of a cancer cell to a
FGFR inhibitor. Non-limiting examples of these mutations are
depicted in Table E. In some embodiments, a FGFR-associated
disorder (e.g., any of the cancers described herein) can have one
or more of the point mutations listed in Table D. Also provided
herein are methods of treating a subject that include identifying a
subject having one or more of the point mutations listed in Table
D, and administering to the identified subject a therapeutically
effective amount of a compound of General Formula I (e.g., any of
the exemplary compounds described herein), or a pharmaceutically
acceptable salt of solvate thereof. Also provided are methods of
treating a subject that include administering to a subject
identified as having one or more of the point mutations listed in
Table D a therapeutically effective amount of a compound of General
Formula I (e.g., any of the exemplary compounds described
herein).
[0294] In some embodiments, provided herein is a method for
treating a subject diagnosed with a FGFR-associated disorder (e.g.,
a FGFR-associated cancer), that include administering to the
subject a therapeutically effective amount of a compound of General
Formula I (e.g., any of the exemplary compounds described herein),
or a pharmaceutically acceptable salt or solvate thereof. For
example, the FGFR-associated cancer can be any of exemplary
FGFR-associated cancers described herein.
[0295] In some embodiments, the compounds of the present invention
are useful for treating a FGFR-associated disease (e.g., a
FGFR-associated cancer) in combination with one or more additional
therapeutic agents or therapies that work by the same or a
different mechanism of action.
[0296] In some embodiments, the additional therapeutic agent(s) is
selected from the group of: receptor tyrosine kinase-targeted
therapeutic agents, including cabozantinib, crizotinib, erlotinib,
gefitinib, imatinib, lapatinib, nilotinib, pazopanib, pertuzumab,
regorafenib, and sunitinib.
[0297] In some embodiments, the additional therapeutic agent(s) is
selected from signal transduction pathway inhibitors, including,
e.g., Ras-Raf-MEK-ERK pathway inhibitors (e.g., sorafenib,
trametinib, or vemurafenib), PI3K-Akt-mTOR-S6K pathway inhibitors
(e.g., everolimus, rapamycin, perifosine, or temsirolimus) and
modulators of the apoptosis pathway (e.g., obataclax).
[0298] In some embodiments, the additional therapeutic agent(s) is
selected from the group of: cytotoxic chemotherapeutics, including,
e.g., arsenic trioxide, bleomycin, cabazitaxel, capecitabine,
carboplatin, cisplatin, cyclophosphamide, cytarabine, dacarbazine,
daunorubicin, docetaxel, doxorubicin, etoposide, fluorouracil,
gemcitabine, irinotecan, lomustine, methotrexate, mitomycin C,
oxaliplatin, paclitaxel, pemetrexed, temozolomide, and
vincristine.
[0299] In some embodiments, the additional therapeutic agent(s) is
selected from the group of angiogenesis-targeted therapies,
including e.g., aflibercept and bevacizumab.
[0300] In some embodiments, the additional therapeutic agent(s) is
selected from the group of immune-targeted agents, e.g., including
aldesleukin, ipilimumab, lambrolizumab, nivolumab, and
sipuleucel-T.
[0301] In some embodiments, the additional therapeutic agent(s) is
selected from agents active against the downstream FGFR pathway,
including, e.g., Ras, MEK, JNK, and p38 kinase inhibitor.
[0302] In some embodiments, the additional therapeutic agent or
therapy is radiotherapy, including, e.g., radioiodide therapy,
external-beam radiation, and radium 223 therapy. In some
embodiments, the additional therapeutic agent(s) includes any one
of the above listed therapies or therapeutic agents which are
standards of care in cancers wherein the cancer has a dysregulation
of a FGFR gene, a FGFR protein, or expression or activity, or level
of the same.
[0303] Methods of detecting dysregulation of a FGFR gene, a FGFR
protein, or expression or activity, or level of the same, include,
e.g., detection of FGFR gene translocations, e.g., using
Fluorescent In Situ Hybridization (FISH) (e.g., the commercially
available kits from Empire Genomics and Cell Signaling
Technology).
[0304] In some embodiments, provided herein is a method of treating
cancer (e.g., a FGFR-associated cancer) in a patient, comprising
administering to said subject a compound of General Formula I
(e.g., any of the exemplary compounds described herein), or a
pharmaceutically acceptable salt or solvate thereof, in combination
with at least one additional therapy or therapeutic agent selected
from radiotherapy (e.g., radioiodide therapy, external-beam
radiation, or radium 223 therapy), cytotoxic chemotherapeutics
(e.g., arsenic trioxide, bleomycin, cabazitaxel, capecitabine,
carboplatin, cisplatin, cyclophosphamide, cytarabine, dacarbazine,
daunorubicin, docetaxel, doxorubicin, etoposide, fluorouracil,
gemcitabine, irinotecan, lomustine, methotrexate, mitomycin C,
oxaliplatin, paclitaxel, pemetrexed, temozolomide, or vincristine);
tyrosine kinase targeted-therapeutics (e.g., afatinib,
cabozantinib, cetuximab, crizotinib, dabrafenib, erlotinib,
gefitinib, imatinib, lapatinib, nilotinib, pazopanib, panitumumab,
pertuzumab, regorafenib, sunitinib, or trastuzumab); FGFR
inhibitors (e.g., ARQ-087, AZD-4547, BGJ398, nintadanib (BIBF
1120), BLU9931, brivanib (BMS-582664), CH5183284, Dovitinib
(TKI258, CHIR258), E-3810, EWMD-2076, JNJ-42756493, lenvatinib
((E7080), LY2874455, Orantinib (TSU-68, SU6668), PD089828,
PD166866, PD173074, Ponatinib (AP-24534), Semaxanib (SU5416),
SSR128129E, SU4984, SU5402, SUN11602), AB1010, BAY 1163877,
Debio-1347, FGF401, FIIN-2, HMPL-453, MK-2461, pazopanib (Votrient,
GW-786034), PD161570, PD173074, PF-477736, PHA-739358 (danusertib),
PRN1371, regorafenib (Stivarga), SPP86, and Tyrphostin AG 1296 and
TAS120; apoptosis modulators and signal transduction inhibitors
(e.g. everolimus, perifosine, rapamycin, sorafenib, temsirolimus,
trametinib, or vemurafenib); immune-targeted therapies (e.g.,
aldesleukin, interferon alfa-2b, ipilimumab, lambrolizumab,
nivolumab, prednisone, or sipuleucel-T); and angiogenesis-targeted
therapies (e.g., aflibercept or bevacizumab), wherein the amount of
the compound of General Formula I (e.g., any of the exemplary
compounds described herein), or a pharmaceutically acceptable salt
or solvate thereof, in combination with the additional therapy or
therapeutic agent, is effective in treating said cancer. These
additional therapeutic agents may be administered with one or more
doses of the compound of General Formula I, or the pharmaceutically
acceptable salt or solvate thereof, as part of the same or separate
dosage forms, via the same or different routes of administration,
and on the same or different administration schedules according to
standard pharmaceutical practice known to one skilled in the art.
In some embodiments, provided herein is a method of treating cancer
(e.g., a FGFR-associated cancer) in a patient, comprising
administering to said subject a compound of General Formula I
(e.g., any of the exemplary compounds described herein), or a
pharmaceutically acceptable salt or solvate thereof, in combination
with at least one additional therapy or therapeutic agent selected
from radiotherapy (e.g., radioiodide therapy, external-beam
radiation, or radium 223 therapy), cytotoxic chemotherapeutics
(e.g., arsenic trioxide, bleomycin, cabazitaxel, capecitabine,
carboplatin, cisplatin, cyclophosphamide, cytarabine, dacarbazine,
daunorubicin, docetaxel, doxorubicin, etoposide, fluorouracil,
gemcitabine, irinotecan, lomustine, methotrexate, mitomycin C,
oxaliplatin, paclitaxel, pemetrexed, temozolomide, or vincristine),
tyrosine kinase targeted-therapeutics (e.g., afatinib,
cabozantinib, cetuximab, crizotinib, dabrafenib, erlotinib,
gefitinib, imatinib, lapatinib, nilotinib, pazopanib, panitumumab,
pertuzumab, regorafenib, or sunitinib), apoptosis modulators and
signal transduction inhibitors (e.g. everolimus, perifosine,
rapamycin, sorafenib, temsirolimus, trametinib, or vemurafenib),
immune-targeted therapies (e.g., aldesleukin, interferon alfa-2b,
ipilimumab, lambrolizumab, nivolumab, prednisone, or sipuleucel-T)
and angiogenesis-targeted therapies (e.g., aflibercept or
bevacizumab), wherein the amount of the compound of General Formula
I (e.g., any of the exemplary compounds described herein), or a
pharmaceutically acceptable salt or solvate thereof, in combination
with the additional therapy or therapeutic agent, is effective in
treating said cancer. These additional therapeutic agents may be
administered with one or more doses of the compound of General
Formula I, or the pharmaceutically acceptable salt or solvate
thereof, as part of the same or separate dosage forms, via the same
or different routes of administration, and on the same or different
administration schedules according to standard pharmaceutical
practice known to one skilled in the art.
[0305] Also provided herein is (i) a pharmaceutical combination for
treating cancer (e.g., a FGFR-associated cancer) in a subject in
need thereof, which comprises (a) a compound of General Formula I
(e.g., any of the exemplary compounds described herein), or a
pharmaceutically acceptable salt or solvate thereof, (b) an
additional therapeutic agent and (c) optionally at least one
pharmaceutically acceptable carrier (e.g., for simultaneous,
separate or sequential use for the treatment of a cancer), wherein
the amounts of the compound of General Formula I, or the
pharmaceutically acceptable salt or solvate thereof and of the
additional therapeutic agent are together effective in treating
said cancer; (ii) a pharmaceutical composition including such a
combination; (iii) the use of such a combination for the
preparation of a medicament for the treatment of cancer (e.g., a
FGFR-associated cancer); and (iv) a commercial package or product
including such a combination as a combined preparation for
simultaneous, separate or sequential use; and to a method of
treatment of cancer (e.g., FGFR-associated cancer) in a subject in
need thereof.
[0306] Also provided are methods of treating a subject identified
or diagnosed as having a FGFR-associated disease (e.g., a
FGFR-associated cancer) (e.g., a subject that has been identified
or diagnosed as having a FGFR-associated disease (e.g., a
FGFR-associated cancer) through the use of a regulatory
agency-approved, e.g., FDA-approved, kit for identifying
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, in a subject or a biopsy sample
from the subject) (e.g., any of the FGFR-associated cancers
described herein or known in the art) that include administering
the subject a therapeutically effective amount of a compound of
General Formula I (e.g., any of the exemplary compounds described
herein), or a pharmaceutically acceptable salt or solvate thereof.
Also provided is a compound of General Formula I (e.g., any of the
exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof, for use in treating a
FGFR-associated disease (e.g., a FGFR-associated cancer) in a
subject identified or diagnosed as having a FGFR-associated disease
(e.g., a FGFR-associated cancer) (e.g., a subject that has been
identified or diagnosed as having a FGFR-associated cancer through
the use of a regulatory agency-approved, e.g., FDA-approved, kit
for identifying dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same, in a subject or a
biopsy sample from the subject) (e.g., any of the FGFR-associated
cancers described herein or known in the art). Also provided is the
use of a compound of General Formula I (e.g., any of the exemplary
compounds described herein), or a pharmaceutically acceptable salt
or solvate thereof, for the manufacture of a medicament for
treating a FGFR-associated disease (e.g., FGFR-associated cancer)
in a subject identified or diagnosed as having a FGFR-associated
disease (e.g., FGFR-associated cancer) (e.g., a subject that has
been identified or diagnosed as having a FGFR-associated cancer
through the use of a regulatory agency-approved, e.g.,
FDA-approved, kit for identifying dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same, in a
subject or a biopsy sample from the subject) (e.g., any of the
FGFR-associated cancers described herein or known in the art).
[0307] Also provided are methods of treating a subject (e.g., a
subject suspected of having a FGFR-associated disease (e.g., a
FGFR-associated cancer), a subject presenting with one or more
symptoms of a FGFR-associated disease (e.g., a FGFR-associated
cancer), or a subject having an elevated risk of developing a
FGFR-associated disease (e.g., FGFR-associated cancer)) that
include performing an assay (e.g., an assay that utilizes next
generation sequencing, immunohistochemistry, or break apart FISH
analysis) (e.g., using a regulatory agency-approved, e.g.,
FDA-approved, kit) on a sample obtained from the subject to
determine whether the subject has dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same, and
administering (e.g., specifically or selectively administering) a
therapeutically effective amount of a compound of General Formula I
(e.g., any of the exemplary compounds described herein), or a
pharmaceutically acceptable salt or solvate thereof, to a subject
determined to have dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or levels of the same. Additional assays,
non-limiting assays that may be used in these methods are described
herein. Additional assays are also known in the art. Also provided
is use of a compound of General Formula I (e.g., any of the
exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof, for use in treating a
FGFR-associated disease (e.g., FGFR-associated cancer) in a subject
identified or diagnosed as having a FGFR-associated disease (e.g.,
a FGFR-associated cancer) through a step of performing an assay
(e.g., an in vitro assay) (e.g., an assay that utilizes next
generation sequencing, immunohistochemistry, or break apart FISH
analysis) (e.g., using a regulatory agency-approved, e.g.,
FDA-approved, kit) on a sample obtained from the subject to
determine whether the subject has dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same,
where the presence of dysregulation of a FGFR gene, a FGFR protein,
or expression or activity, or level of the same, identifies that
the subject has a FGFR-associated disorder (e.g., FGFR-associated
cancer). Also provided is the use of a compound of General Formula
I (e.g., any of the exemplary compounds described herein), or a
pharmaceutically acceptable salt or solvate thereof, for the
manufacture of a medicament for treating a FGFR-associated disease
(e.g., FGFR-associated cancer) in a subject identified or diagnosed
as having a FGFR-associated disease (e.g., FGFR-associated cancer)
through a step of performing an assay (e.g., an in vitro assay)
(e.g., an assay that utilizes next generation sequencing,
immunohistochemistry, or break apart FISH analysis) (e.g., using a
regulatory agency-approved, e.g., FDA-approved, kit) on a sample
obtained from the subject to determine whether the subject has
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, where the presence of dysregulation
of a FGFR gene, a FGFR protein, or expression or activity, or level
of the same, identifies that the subject has a FGFR-associated
disease (e.g., FGFR-associated cancer). Some embodiments of any of
the methods or uses described herein further include recording in
the subject's clinical record (e.g., a computer readable medium)
that the subject determined to have dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same,
through the performance of the assay, should be administered a
compound of General Formula I (e.g., any of the exemplary compounds
described herein), or a pharmaceutically acceptable salt or solvate
thereof.
[0308] In some embodiments of any of the methods or uses described
herein, the subject has been identified or diagnosed as having a
cancer with dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same (e.g., as determined
using a regulatory agency-approved, e.g., FDA-approved, assay or
kit). In some embodiments of any of the methods or uses described
herein, the subject has a tumor that is positive for dysregulation
of a FGFR gene, a FGFR protein, or expression or activity, or level
of the same (e.g., as determined using a regulatory agency-approved
assay or kit). In some embodiments of any of the methods or uses
described herein, the subject can be a subject with a tumor(s) that
is positive for dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same (e.g., identified as
positive using a regulatory agency-approved, e.g., FDA-approved,
assay or kit). In some embodiments of any of the methods or uses
described herein, the subject can be a subject whose tumors have
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or a level of the same (e.g., where the tumor is
identified as such using a regulatory agency-approved, e.g.,
FDA-approved, kit or assay). In some embodiments of any of the
methods or uses described herein, the subject is suspected of
having a FGFR-associated cancer. In some embodiments of any of the
methods or uses described herein, the subject has a clinical record
indicating that the subject has a tumor that has dysregulation of a
FGFR gene, a FGFR protein, or expression or activity, or level of
the same (and optionally the clinical record indicates that the
subject should be treated with any of the compositions provided
herein).
[0309] Also provided are methods of treating a subject that include
administering a therapeutically effective amount of a compound of
General Formula I (e.g., any of the exemplary compounds described
herein), or a pharmaceutically acceptable salt or solvate thereof,
to a subject having a clinical record that indicates that the
subject has dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same. Also provided is the
use of a compound of General Formula I (e.g., any of the exemplary
compounds described herein), or a pharmaceutically acceptable salt
or solvate thereof, for the manufacture of a medicament for
treating a FGFR-associated disease (e.g., FGFR-associated cancer)
in a subject having a clinical record that indicates that the
subject has dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same. Also provided is the
use of a compound of General Formula I (e.g., any of the exemplary
compounds described herein), or a pharmaceutically acceptable salt
or solvate thereof, for the manufacture of a medicament for
treating a FGFR-associated disease (e.g., a FGFR-associated cancer)
in a subject having a clinical record that indicates that the
subject has dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same. Some embodiments of
these methods and uses can further include: a step of performing an
assay (e.g., an in vitro assay) (e.g., an assay that utilizes next
generation sequencing, immunohistochemistry, or break apart FISH
analysis) (e.g., using a regulatory agency-approved, e.g.,
FDA-approved, kit) on a sample obtained from the subject to
determine whether the subject has dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same, and
recording information in a subject's clinical file (e.g., a
computer-readable medium) that the subject has been identified to
have dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same.
[0310] Also provided are methods (e.g., in vitro methods) of
selecting a treatment for a subject that include selecting a
treatment including administration of a therapeutically effective
amount of a compound of General Formula I (e.g., any of the
exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof, for a subject identified or
diagnosed as having a FGFR-associated disease (e.g., a
FGFR-associated cancer) (e.g., a subject that has been identified
or diagnosed as having a FGFR-associated cancer through the use of
a regulatory agency-approved, e.g., FDA-approved, kit for
identifying dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or level of the same, in a subject or a
biopsy sample from the subject) (e.g., any of the FGFR-associated
cancers described herein or known in the art). Some embodiments can
further include administering the selected treatment to the subject
identified or diagnosed as having a FGFR-associated disease (e.g.,
a FGFR-associated cancer). Some embodiments can further include a
step of performing an assay (e.g., an in vitro assay) (e.g., an
assay that utilizes next generation sequencing,
immunohistochemistry, or break apart FISH analysis) (e.g., using a
regulatory agency-approved, e.g., FDA-approved, kit) on a sample
obtained from the subject to determine whether the subject has
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, and identifying or diagnosing a
subject determined to have dysregulation of a FGFR gene, a FGFR
protein, or expression or activity, or level of the same, as having
a FGFR-associated disease (e.g., a FGFR-associated cancer).
[0311] Also provided are methods of selecting a treatment for a
subject that include administration of a therapeutically effective
amount of a compound of General Formula I (e.g., any of the
exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof, wherein the methods include a
step of performing an assay (e.g., an in vitro assay) (e.g., an
assay that utilizes next generation sequencing,
immunohistochemistry, or break apart FISH analysis) (e.g., using a
regulatory agency-approved, e.g., FDA-approved, kit) on a sample
obtained from the subject to determine whether the subject has
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, and identifying or diagnosing a
subject determined to have dysregulation of a FGFR gene, a FGFR
protein, or expression or activity, or level of the same, as having
a FGFR-associated cancer, and selecting a therapeutic treatment
including administration of a therapeutically effective amount of a
compound of General Formula I (e.g., any of the exemplary compounds
described herein), or a pharmaceutically acceptable salt or solvate
thereof, for the subject identified or diagnosed as having a
FGFR-associated disease (e.g., a FGFR-associated cancer). Some
embodiments further include administering the selected treatment to
the subject identified or diagnosed as having a FGFR-associated
disorder (e.g., a FGFR-associated cancer).
[0312] Also provided are methods of selecting a subject for
treatment including administration of a therapeutically effective
amount of a compound of General Formula I (e.g., any of the
exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof, that include selecting,
identifying, or diagnosing a subject having a FGFR-associated
disorder (e.g., a FGFR-associated cancer), and selecting the
subject for treatment including administration of a therapeutically
effective amount of a compound of General Formula I (e.g., any of
the exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof. In some embodiments,
identifying or diagnosing a subject as having a FGFR-associated
disease (e.g., an FGFR-associated cancer) can include a step of
performing an assay (e.g., an in vitro assay) (e.g., an assay that
utilizes next generation sequencing, immunohistochemistry, or break
apart FISH analysis) (e.g., using a regulatory agency-approved,
e.g., FDA-approved, kit) on a sample obtained from the subject to
determine whether the subject has dysregulation of a FGFR gene, a
FGFR protein, or expression or activity, or level of the same, and
identifying or diagnosing a subject determined to have
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, as having a FGFR-associated
disorder (e.g., a FGFR-associated cancer). In some embodiments, the
selecting a treatment can be used as part of a clinical study that
includes administration of various treatments of an FGFR-associated
disorder (e.g., a FGFR-associated cancer).
[0313] In some embodiments of any of the methods or uses described
herein, an assay used determine whether the subject has
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity, or level of the same, using a sample (e.g., a biological
sample or a biopsy sample (e.g., a paraffin-embedded biopsy sample)
from a subject (e.g., a subject suspected of having a
FGFR-associated disease (e.g., a FGFR-associated cancer), a subject
having one or more symptoms of a FGFR-associated disease (e.g., a
FGFR-associated cancer), and/or a subject that has an increased
risk of developing a FGFR-associated disease (e.g., a
FGFR-associated cancer)) can include, for example, next generation
sequencing, immunohistochemistry, fluorescence microscopy, break
apart FISH analysis, Southern blotting, Western blotting, FACS
analysis, Northern blotting, and PCR-based amplification (e.g.,
RT-PCR). As is well-known in the art, the assays are typically
performed, e.g., with at least one labelled nucleic acid probe or
at least one labelled antibody or antigen-binding fragment thereof.
Assays can utilize other detection methods known in the art for
detecting dysregulation of a FGFR gene, a FGFR protein, or
expression or activity, or levels of the same (see, e.g., the
references cited herein).
[0314] Exemplary assays for detecting dysregulation of a FGFR gene,
a FGFR protein, or expression or activity, or levels of the same
are commercially available, e.g., FGFR Pathway Mutation PCR Array
(Qiagen), HTG Edge FGFR Expression Assay (HTG Molecular
Diagnostics), HTScan.RTM. FGF Receptor 1 Kinase Assay Kit (Cell
Signaling Technology), Vysis LSI IGH/FGFR3 Dual Color, Dual Fusion
Translocation Probe (Abbott Molecular), FGFR1 FISH Probe (Empire
Genomics), FGFR1 FISH (Sonic Genomics), FISH IGH/FGFR3 (Quest
Diagnostics), FGFR1 (8p11) [RUO] (Leica Biosystems), FGFR1 Break
Apart FISH Probe (Empire Genomics), FGFR2/CEN10p FISH Probe (Abnova
Corporation), FGFR2 (10q26) [ASR](Leica Biosystems), Anti-FGFR-4
(IN), Z-FISH (AnaSpec), ZytoLight.RTM. SPEC FGFR2 Break Apart Probe
(Bio-Optica), FGFR3 (4p16.3) (ZytoVision), and ZytoLight.RTM. SPEC
FGFR3/CEN4 Dual Color Probe (ZytoVision). Additional assays for
detecting dysregulation of a FGFR gene, a FGFR protein, or
expression or activity or levels of the same are known in the
art.
[0315] Also provided are methods of increasing the time of
remission of a FGFR-associated cancer in a patient that include (a)
selecting, identifying, or diagnosing a patient as having a
FGFR-associated cancer (e.g., any of the FGFR-associated cancers
described herein), and (b) administering a therapeutically
effective amount of a compound of General Formula I (e.g., any of
the exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof. Also provided are methods of
increasing the time of remission of a FGFR-associated cancer in a
patient that include administering a therapeutically effective
amount of a compound of General Formula I (e.g., any of the
exemplary compounds described herein), or a pharmaceutically
acceptable salt or solvate thereof to a patient having a
FGFR-associated cancer (e.g., any of the exemplary FGFR-associated
cancers described herein). In some examples of any of the methods
of increasing the time of remission of a FGFR-associated cancer in
a patient, the increase in the time of remission is compared to a
control patient (e.g., a patient or a population of patients having
the same or a similar type of FGFR-associated cancer). In some
examples, the patient is not yet in remission. In other examples,
the patient is already in remission. In some examples, the increase
in remission is a statistically significant increase. In some
examples, the increase in the time of remission is about 1 day to
about 10 years, about 9.5 years, about 9 years, about 8.5 years,
about 8 years, about 7.5 years, about 7 years, about 6.5 years,
about 6 years, about 5.5 years, about 5 years, about 4.5 years,
about 4 years, about 3.5 years, about 3 years, about 2.5 years,
about 2 years, about 1.5 years, about 1 year, about 10 months,
about 8 months, about 6 months, about 4 months, about 2 months,
about 1 month, or about 2 weeks; about 2 weeks to about 10 years,
about 9.5 years, about 9 years, about 8.5 years, about 8 years,
about 7.5 years, about 7 years, about 6.5 years, about 6 years,
about 5.5 years, about 5 years, about 4.5 years, about 4 years,
about 3.5 years, about 3 years, about 2.5 years, about 2 years,
about 1.5 years, about 1 year, about 10 months, about 8 months,
about 6 months, about 4 months, about 2 months, or about 1 month;
about 1 month to about 10 years, about 9.5 years, about 9 years,
about 8.5 years, about 8 years, about 7.5 years, about 7 years,
about 6.5 years, about 6 years, about 5.5 years, about 5 years,
about 4.5 years, about 4 years, about 3.5 years, about 3 years,
about 2.5 years, about 2 years, about 1.5 years, about 1 year,
about 10 months, about 8 months, about 6 months, about 4 months, or
about 2 months; about 2 month to about 10 years, about 9.5 years,
about 9 years, about 8.5 years, about 8 years, about 7.5 years,
about 7 years, about 6.5 years, about 6 years, about 5.5 years,
about 5 years, about 4.5 years, about 4 years, about 3.5 years,
about 3 years, about 2.5 years, about 2 years, about 1.5 years,
about 1 year, about 10 months, about 8 months, about 6 months, or
about 4 months; about 4 month to about 10 years, about 9.5 years,
about 9 years, about 8.5 years, about 8 years, about 7.5 years,
about 7 years, about 6.5 years, about 6 years, about 5.5 years,
about 5 years, about 4.5 years, about 4 years, about 3.5 years,
about 3 years, about 2.5 years, about 2 years, about 1.5 years,
about 1 year, about 10 months, about 8 months, or about 6 months;
about 6 month to about 10 years, about 9.5 years, about 9 years,
about 8.5 years, about 8 years, about 7.5 years, about 7 years,
about 6.5 years, about 6 years, about 5.5 years, about 5 years,
about 4.5 years, about 4 years, about 3.5 years, about 3 years,
about 2.5 years, about 2 years, about 1.5 years, about 1 year,
about 10 months, or about 8 months; about 8 month to about 10
years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, about 5 years, about 4.5 years, about 4
years, about 3.5 years, about 3 years, about 2.5 years, about 2
years, about 1.5 years, about 1 year, or about 10 months; about 10
month to about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, about 5 years, about 4.5
years, about 4 years, about 3.5 years, about 3 years, about 2.5
years, about 2 years, about 1.5 years, or about 1 year; about 1
year to about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, about 5 years, about 4.5
years, about 4 years, about 3.5 years, about 3 years, about 2.5
years, about 2 years, or about 1.5 years; about 1.5 years to about
10 years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, about 5 years, about 4.5 years, about 4
years, about 3.5 years, about 3 years, about 2.5 years, to about 2
years; about 2 years to about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, about 6.5 years, about 6 years, about 5.5 years, about 5
years, about 4.5 years, about 4 years, about 3.5 years, about 3
years, or about 2.5 years; about 2.5 years to about 10 years, about
9.5 years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, about 5.5
years, about 5 years, about 4.5 years, about 4 years, about 3.5
years, or about 3 years; about 3 years to about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, about 5.5
years, about 5 years, about 4.5 years, about 4 years, or about 3.5
years; about 3.5 years to about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, about 6.5 years, about 6 years, about 5.5 years, about 5
years, about 4.5 years, or about 4 years; about 4 years to about 10
years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, about 5 years, or about 4.5 years; about
4.5 years to about 10 years, about 9.5 years, about 9 years, about
8.5 years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, or about 5 years; about 5
years to about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, or about 5.5 years; about 5.5 years to about
10 years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, or about 6
years; about 6 years to about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, or about 6.5 years; about 6.5 years to about 10 years, about
9.5 years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, or about 7 years; about 7 years to about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, or about 7.5
years; about 7.5 years to about 10 years, about 9.5 years, about 9
years, about 8.5 years, or about 8 years; about 8 years to about 10
years, about 9.5 years, about 9 years, or about 8.5 years; about
8.5 years to about 10 years, about 9.5 years, or about 9 years;
about 9 years to about 10 years or about 9.5 years; or about 9.5
years to about 10 years (e.g., compared to a control patient, e.g.,
a patient or a population of patients having the same or a similar
type of FGFR-associated cancer).
[0316] Also provided is a compound of General Formula I or
pharmaceutically acceptable salt or solvate thereof for use in
increasing the time of remission of a FGFR-associated cancer in a
patient. Also provided is the use of a compound of General Formula
I or a pharmaceutically acceptable salt or solvate thereof in the
manufacture of a medicament for increasing the time of remission of
a FGFR-associated cancer in a patient.
[0317] Methods for determining whether or not a patient is in
remission are known by those skilled in the art. For example, a PET
scan, MRI, CT scan, ultrasound, and X-ray of the patient's body may
be obtained, and such data can be used to determine whether or not
a patient is in remission. In some examples, diagnostic tests can
be performed on samples from a patient (e.g., a blood sample or a
biopsy) to determine whether or not the patient is still in
remission.
Also provided are methods of increasing the time of survival of a
patient having a FGFR-associated cancer that include: selecting,
diagnosing, or identifying a patient as having a FGFR-associated
cancer; and administering to a subject selected, diagnosed, or
identified as having a FGFR-associated cancer a therapeutically
effective amount of a compound of General Formula I or a
pharmaceutically acceptable salt or solvate thereof. Also provided
are methods of increasing the time of survival of a patient having
a FGFR-associated cancer that include administering to a subject
having a FGFR-associated cancer a therapeutically effective amount
of a compound of General Formula I or a pharmaceutically acceptable
salt or solvate thereof. In some embodiments of any of the methods
of increasing the time of survival of a subject having a
FGFR-associated cancer, the increase in the time of survival is
compared to a control patient (e.g., a patient or a population of
patients having the same or a similar type of FGFR-associated
cancer). In some examples, the patient can have an early stage of a
FGFR-associated cancer (e.g., Stage 1 or 2). In some embodiments,
the patient can have a late stage of a FGFR-associated cancer
(e.g., Stage 3 or 4). In some examples, the increase in the time of
survival is a statistically significant increase. In some examples,
the increase in the time of survival is about 1 day to about 40
years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, about 5.5
years, about 5 years, about 4.5 years, about 4 years, about 3.5
years, about 3 years, about 2.5 years, about 2 years, about 1.5
years, about 1 year, about 10 months, about 8 months, about 6
months, about 4 months, about 2 months, about 1 month, or about 2
weeks; about 2 weeks to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, about 5 years, about 4.5
years, about 4 years, about 3.5 years, about 3 years, about 2.5
years, about 2 years, about 1.5 years, about 1 year, about 10
months, about 8 months, about 6 months, about 4 months, about 2
months, or about 1 month; about 1 month to about 40 years, about 38
years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, about 12 years, about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, about 6.5 years, about 6 years, about 5.5 years, about 5
years, about 4.5 years, about 4 years, about 3.5 years, about 3
years, about 2.5 years, about 2 years, about 1.5 years, about 1
year, about 10 months, about 8 months, about 6 months, about 4
months, or about 2 months; about 2 months to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, about 12 years, about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, about 6.5 years, about 6 years, about 5.5 years, about 5
years, about 4.5 years, about 4 years, about 3.5 years, about 3
years, about 2.5 years, about 2 years, about 1.5 years, about 1
year, about 10 months, about 8 months, about 6 months, or about 4
months; about 4 months to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, about 5 years, about 4.5
years, about 4 years, about 3.5 years, about 3 years, about 2.5
years, about 2 years, about 1.5 years, about 1 year, about 10
months, about 8 months, or about 6 months; about 6 months to about
40 years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, about 5.5
years, about 5 years, about 4.5 years, about 4 years, about 3.5
years, about 3 years, about 2.5 years, about 2 years, about 1.5
years, about 1 year, about 10 months, or about 8 months; about 8
months to about 40 years, about 38 years, about 36 years, about 34
years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, about 20 years, about 18
years, about 16 years, about 14 years, about 12 years, about 10
years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, about 5 years, about 4.5 years, about 4
years, about 3.5 years, about 3 years, about 2.5 years, about 2
years, about 1.5 years, about 1 year, or about 10 months; about 10
months to about 40 years, about 38 years, about 36 years, about 34
years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, about 20 years, about 18
years, about 16 years, about 14 years, about 12 years, about 10
years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, about 5 years, about 4.5 years, about 4
years, about 3.5 years, about 3 years, about 2.5 years, about 2
years, about 1.5 years, or about 1 year; about 1 year to about 40
years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, about 5.5
years, about 5 years, about 4.5 years, about 4 years, about 3.5
years, about 3 years, about 2.5 years, about 2 years, or about 1.5
years; about 1.5 year to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, about 5 years, about 4.5
years, about 4 years, about 3.5 years, about 3 years, about 2.5
years, or about 2 years; about 2 year to about 40 years, about 38
years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, about 12 years, about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, about 6.5 years, about 6 years, about 5.5 years, about 5
years, about 4.5 years, about 4 years, about 3.5 years, about 3
years, or about 2.5 years; about 2.5 year to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, about 12 years, about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, about 6.5 years, about 6 years, about 5.5 years, about 5
years, about 4.5 years, about 4 years, about 3.5 years, or about 3
years; about 3 year to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, about 6 years, about 5.5 years, about 5 years, about 4.5
years, about 4 years, or about 3.5 years; about 3.5 years to about
40 years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, about 5.5
years, about 5 years, about 4.5 years, or about 4 years; about 4
years to about 40 years, about 38 years, about 36 years, about 34
years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, about 20 years, about 18
years, about 16 years, about 14 years, about 12 years, about 10
years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, about 5 years, or about 4.5 years; about
4.5 years to about 40 years, about 38 years, about 36 years, about
34 years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, about 20 years, about 18
years, about 16 years, about 14 years, about 12 years, about 10
years, about 9.5 years, about 9 years, about 8.5 years, about 8
years, about 7.5 years, about 7 years, about 6.5 years, about 6
years, about 5.5 years, or about 5 years; about 5 years to about 40
years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, about 9.5
years, about 9 years, about 8.5 years, about 8 years, about 7.5
years, about 7 years, about 6.5 years, about 6 years, or about 5.5
years; about 5.5 years to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, about 7.5 years, about 7 years, about 6.5
years, or about 6 years; about 6 years to about 40 years, about 38
years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, about 12 years, about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, about 7
years, or about 6.5 years; about 6.5 years to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, about 12 years, about 10 years, about 9.5 years, about 9
years, about 8.5 years, about 8 years, about 7.5 years, or about 7
years, about 7 years to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, about 10 years, about 9.5 years, about 9 years, about 8.5
years, about 8 years, or about 7.5 years; about 7.5 years to about
40 years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, about 9.5
years, about 9 years, about 8.5 years, or about 8 years; about 8
years to about 40 years, about 38 years, about 36 years, about 34
years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, about 20 years, about 18
years, about 16 years, about 14 years, about 12 years, about 10
years, about 9.5 years, about 9 years, or about 8.5 years; about
8.5 years to about 40 years, about 38 years, about 36 years, about
34 years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, about 20 years, about 18
years, about 16 years, about 14 years, about 12 years, about 10
years, about 9.5 years, or about 9 years; about 9 years to about 40
years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, about 18 years, about 16
years, about 14 years, about 12 years, about 10 years, or about 9.5
years; about 9.5 years to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, about 16 years, about 14 years, about 12
years, or about 10 years; about 10 years to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, about 14
years, or about 12 years; about 12 years to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, about 30
years, about 28 years, about 26 years, about 24 years, about 22
years, about 20 years, about 18 years, about 16 years, or about 14
years; about 14 years to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, about 26 years, about 24 years, about 22 years, about 20
years, about 18 years, or about 16 years; about 16 years to about
40 years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, about 24
years, about 22 years, about 20 years, or about 18 years; about 18
years to about 40 years, about 38 years, about 36 years, about 34
years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, about 22 years, or about 20 years; about 20
years to about 40 years, about 38 years, about 36 years, about 34
years, about 32 years, about 30 years, about 28 years, about 26
years, about 24 years, or about 22 years; about 22 years to about
40 years, about 38 years, about 36 years, about 34 years, about 32
years, about 30 years, about 28 years, about 26 years, or about 24
years; about 24 years to about 40 years, about 38 years, about 36
years, about 34 years, about 32 years, about 30 years, about 28
years, or about 26 years; about 26 years to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, about 30
years, or about 28 years; about 28 years to about 40 years, about
38 years, about 36 years, about 34 years, about 32 years, or about
30 years; about 30 years to about 40 years, about 38 years, about
36 years, about 34 years, or about 32 years; about 32 years to
about 40 years, about 38 years, about 36 years, or about 34 years;
about 34 years to about 40 years, about 38 years, or about 36
years; about 36 years to about 40 years or about 38 years; or about
38 years to about 40 years (e.g., compared to a control patient,
e.g., a patient or a population of patients having the same or a
similar type of FGFR-associated cancer).
[0319] Also provided is the use of a compound of General Formula I
or a pharmaceutically acceptable salt or solvate thereof for
increasing the time of survival of a patient having a
FGFR-associated cancer. Also provided is the use of a compound of
General Formula I or a pharmaceutically acceptable salt or solvate
thereof for the manufacture of a medicament for increasing the time
of survival of a patient having a FGFR-associated cancer.
[0320] Also provided are methods of decreasing the risk of
developing a metastasis or an additional metastasis in a patient
having a FGFR-associated cancer that include: selecting,
identifying, or diagnosing a patient as having a FGFR-associated
cancer, and administering a therapeutically effective amount of a
compound of General Formula I or a pharmaceutically acceptable salt
or solvate thereof to the patient selected, identified, or
diagnosed as having a FGFR-associated cancer. Also provided are
methods of decreasing the risk of developing a metastasis or an
additional metastasis in a patient having a FGFR-associated cancer
that includes administering a therapeutically effective amount of a
compound of General Formula I or a pharmaceutically acceptable salt
or solvent thereof to a patient having a FGFR-associated cancer.
The decrease in the risk of developing a metastasis or an
additional metastasis in a patient having a FGFR-associated cancer
can be compared to the risk of developing a metastasis or an
additional metastasis in the patient prior to treatment, or as
compared to a patient or a population of patients having a similar
or the same FGFR-associated cancer that has received no treatment
or a different treatment. The decrease in the risk of developing a
metastasis or an additional metastasis can be about 1% to about
99%, about 95%, about 90%, about 85%, about 80%, about 75%, about
70%, about 65%, about 60%, about 55%, about 50%, about 45%, about
40%, about 35%, about 30%, about 25%, about 20%, about 15%, about
10%, or about 5%; about 5% to about 99%, about 95%, about 90%,
about 85%, about 80%, about 75%, about 70%, about 65%, about 60%,
about 55%, about 50%, about 45%, about 40%, about 35%, about 30%,
about 25%, about 20%, about 15%, or about 10%; about 10% to about
99%, about 95%, about 90%, about 85%, about 80%, about 75%, about
70%, about 65%, about 60%, about 55%, about 50%, about 45%, about
40%, about 35%, about 30%, about 25%, about 20%, or about 15%;
about 15% to about 99%, about 95%, about 90%, about 85%, about 80%,
about 75%, about 70%, about 65%, about 60%, about 55%, about 50%,
about 45%, about 40%, about 35%, about 30%, about 25%, or about
20%; about 20% to about 99%, about 95%, about 90%, about 85%, about
80%, about 75%, about 70%, about 65%, about 60%, about 55%, about
50%, about 45%, about 40%, about 35%, about 30%, or about 25%;
about 25% to about 99%, about 95%, about 90%, about 85%, about 80%,
about 75%, about 70%, about 65%, about 60%, about 55%, au bout 45%,
about 40%, about 35%, or about 30%; about 30% to about 99%, about
95%, about 90%, about 85%, about 80%, about 75%, about 70%, about
65%, about 60%, about 55%, about 50%, about 45%, about 40%, or
about 35%; about 35% to about 99%, about 95%, about 90%, about 85%,
about 80%, about 75%, about 70%, about 65%, about 60%, about 55%,
about 50%, about 45%, or about 40%; about 40% to about 99%, about
95%, about 90%, about 85%, about 80%, about 75%, about 70%, about
65%, about 60%, about 55%, about 50%, or about 45%; about 45% to
about 99%, about 95%, about 90%, about 85%, about 80%, about 75%,
about 70%, about 65%, about 60%, about 55%, or about 50%; about 50%
to about 99%, about 95%, about 90%, about 85%, about 80%, about
75%, about 70%, about 65%, about 60%, or about 55%; about 55% to
about 99%, about 95%, about 90%, about 85%, about 80%, about 75%,
about 70%, about 65%, or about 60%; about 60% to about 99%, about
95%, about 90%, about 85%, about 80%, about 75%, about 70%, or
about 65%; about 65% to about 99%, about 95%, about 90%, about 85%,
about 80%, about 75%, or about 70%; about 70% to about 99%, about
95%, about 90%, about 85%, about 80%, or about 75%; about 75% to
about 99%, about 95%, about 90%, about 85%, or about 80%; about 80%
to about 99%, about 95%, about 90%, or about 85%; about 85% to
about 99%, about 95%, or about 90%; about 90% to about 99% or about
90%; or about 95% to about 99% as compared to the risk of
developing a metastasis or an additional metastasis in the patient
prior to treatment, or as compared to a patient or a population of
patients having a similar or the same FGFR-associated cancer that
has received no treatment or a different treatment.
[0321] In some examples, the risk of developing a metastasis or an
additional metastasis is over about 2 weeks, 1 month, 1.5 months, 2
months, 2.5 months, 3 months, 3.5 months, 4 months, 4.5 months, 5
months, 5.5 months, 6 months, 6.5 months, 7 months, 7.5 months, 8
months, 8.5 months, 9 months, 9.5 months, 10 months, 10.5 months,
11 months, 11.5 months, 12 months, 1.5 years, 2 years, 2.5 years, 3
years, 3.5 years, 4 years, 4.5 years, 5 years, 5.5 years, 6 years,
6.5 years, 7 years, 7.5 years, 8 years, 8.5 years, 9 years, 9.5
years, or 10 years.
[0322] Also provided is the use of a compound of General Formula I
or a pharmaceutically acceptable salt or solvate thereof for
decreasing the risk of developing a metastasis or an additional
metastasis in a patient having a FGFR-associated cancer. Also
provided is the use of a compound of General Formula I or a
pharmaceutically acceptable salt or solvate thereof for the
manufacture of a medicament for decreasing the risk of developing a
metastasis or an additional metastasis in a patient having a
FGFR-associated cancer.
[0323] Also provided are methods of increasing sensitivity of a
resistant cancer cell to an anti-cancer drug that include:
selecting, identifying, or diagnosing a patient as having a
resistant cancer cell (e.g., a resistant FGFR-associated cancer
cell, e.g., a cancer cell identified as having one or more of the
point mutations listed in Table E), and administering to the
selected, identified, or diagnosed subject a therapeutically
effective amount of a compound of General Formula I or a
pharmaceutically acceptable salt or solvate thereof. Also provided
are methods of increasing sensitivity of a resistant cancer cell to
an anti-cancer drug that include administering to a patient having
a resistant cancer cell to an anti-cancer drug a therapeutically
effective amount of a compound of General Formula I or a
pharmaceutically acceptable salt or solvate thereof. Some
embodiments of any of these methods further include administering
the anti-cancer drug to the patient. In such examples, the
anti-cancer drug can be co-administered with the compound of
General Formula I or a pharmaceutically acceptable salt or solvate
thereof. In some examples, the anti-cancer drug can be administered
at substantially the same time as the compound of General Formula I
or a pharmaceutically acceptable salt or solvate thereof. In some
examples, a first dose of the compound of General Formula I is
administered prior to the first dose of the anti-cancer compound.
In some examples, a first dose of the anti-cancer compound is
administered prior to the first dose of the compound of General
Formula I or a pharmaceutically acceptable salt or solvate thereof.
In some examples, the increase in the sensitivity of the resistant
cancer cell to the anti-cancer drug can result in a decrease in the
rate of growth and/or proliferation of the resistant cancer cell
when contacted with the anti-cancer drug and at least one of the
compounds described herein, of between about 1% to about 100%, 95%,
90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%,
25%, 20%, 15%, 10%, or 5%; about 2% to about 100%, 95%, 90%, 85%,
80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%,
15%, 10%, or 5%; about 3% to about 100%, 95%, 90%, 85%, 80%, 75%,
70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, or
5%; about 5% to about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%,
55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, or 10%; about 5% to
about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%,
40%, 35%, 30%, 25%, 20%, 15%, 10%, or 5%; about 5% to about 100%,
95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%,
30%, 25%, 20%, 15%, or 10%; about 10% to about 100%, 95%, 90%, 85%,
80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%,
15%, 10%, or 5%; about 5% to about 100%, 95%, 90%, 85%, 80%, 75%,
70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, or 15%;
about 15% to about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%,
55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, or 5%; about 5%
to about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%,
45%, 40%, 35%, 30%, 25%, or 20%; about 20% to about 100%, 95%, 90%,
85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%,
20%, 15%, 10%, or 5%; about 5% to about 100%, 95%, 90%, 85%, 80%,
75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, or 25%; about 25%
to about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%,
45%, 40%, 35%, 30%, 25%, or 20%; about 20% to about 100%, 95%, 90%,
85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%,
20%, 15%, 10%, or 5%; about 5% to about 100%, 95%, 90%, 85%, 80%,
75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, or 30%; about 30% to
about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%,
40%, or 35%; about 35% to 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%,
60%, 55%, 50%, 45%, or 40%; about 40% to about 100%, 95%, 90%, 85%,
80%, 75%, 70%, 65%, 60%, 55%, 50%, or 45%; about 45% to about 100%,
95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, or 50%; about 50% to
about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, or 55%; about
55% to about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, or 60%; about
60% to about 100%, 95%, 90%, 85%, 80%, 75%, 70%, or 65%; about 65%
to about 100%, 95%, 90%, 85%, 80%, 75%, or 70%; about 70% to about
100%, 95%, 90%, 85%, 80%, or 75%; about 75% to about 100%, 95%,
90%, 85%, or 80%; about 80% to about 100%, 95%, 90%, or 85%; about
85% to about 100%, 95%, or 90%; about 90% to about 100% or 95%; or
about 95% to about 100%, as compared to the rate of growth and/or
proliferation of a resistant cancer cell when contacted with the
anti-cancer drug alone.
[0324] A method of treating an angiogenesis-related disorder (e.g.,
any of the angiogenesis-related disorders described herein or known
in the art) in a patient that include: identifying, selecting, or
diagnosing a angiogenesis-related disorder in a patient, and
administering to the identified, selected, or diagnosed patient
with a therapeutically effective amount of a compound of General
Formula I or a pharmaceutically acceptable salt or solvate thereof.
A method of treating an angiogenesis-related disorder in a patient
that includes administering a therapeutically effective amount of a
compound of General Formula I or a pharmaceutically acceptable salt
or solvent thereof to a patient having an angiogenesis-related
disorder. In some examples, the treating can result in a decrease
in the diameter of a blood vessel and/or a decrease in the number
of blood vessels in a tissue in need of a reduction in the number
of blood vessels (e.g., as compared to the diameter of the blood
vessel and/or the number of blood vessels in the tissue in the
patient prior to treatment). In some examples the methods can
result in, e.g., a decrease in the diameter of a blood vessel of
about 1% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%,
30%, 25%, 20%, 15%, 10%, or 5%; about 2% to about 80%, 75%, 70%,
65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, or 5%;
about 3% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%,
30%, 25%, 20%, 15%, 10%, or 5%; about 5% to about 80%, 75%, 70%,
65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, or 10%;
about 10% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%,
35%, 30%, 25%, 20%, or 15%; about 15% to about 80%, 75%, 70%, 65%,
60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, or 20%; about 20% to about
80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, or 25%;
about 25% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%,
35%, or 30%; about 30% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%,
45%, 40%, or 35%; about 35% to about 80%, 75%, 70%, 65%, 60%, 55%,
50%, 45%, or 40%; about 40% to about 80%, 75%, 70%, 65%, 60%, 55%,
50%, or 45%; about 45% to about 80%, 75%, 70%, 65%, 60%, 55%, or
50%; about 50% to about 80%, 75%, 70%, 65%, 60%, or 55%; about 55%
to about 80%, 75%, 70%, 65%, or 60%; about 60% to about 80%, 75%,
70%, or 65%; about 65% to about 80%, 75%, or 70%; about 70% to
about 80% or 75%, or about 75% to about 80% (e.g., as compared to
the diameter of the blood vessel in the patient prior to
treatment). In some examples the methods can result in, e.g., a
decrease in the number of blood vessels in a tissue in need of a
reduction in the number of blood vessels of about 5% to about 80%,
75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, or
10%; about 10% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%,
40%, 35%, 30%, 25%, 20%, or 15%; about 15% to about 80%, 75%, 70%,
65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, or 20%; about 20% or
about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, or
25%; about 25% to about 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%,
40%, 35%, or 30%; about 30% to about about 80%, 75%, 70%, 65%, 60%,
55%, 50%, 45%, 40%, or 35%; about 35% to about 80%, 75%, 70%, 65%,
60%, 55%, 50%, 45%, or 40%; about 40% to about 80%, 75%, 70%, 65%,
60%, 55%, 50%, or 45%; about 45% to about 80%, 75%, 70%, 65%, 60%,
55%, or 50%; about 50% to about 80%, 75%, 70%, 65%, 60%, or 55%;
about 55% to about 80%, 75%, 70%, 65%, or 60%; about 60% to about
80%, 75%, 70%, or 65%; about 65% to about 80%, 75%, or 70%; about
70% to about 80% or 75%; or about 75% to about 80% (e.g., as
compared to the diameter of the blood vessel and/or the number of
blood vessels in the tissue in the patient prior to treatment).
These methods can also result in a decrease in the rate of
formation of new blood vessels in a tissue in need thereof in a
patient having an angiogenesis-related disorder (e.g., as compared
to the rate of formation of new blood vessels in the tissue in the
patient prior to treatment, or the rate of formation of new blood
vessels in a patient or a population of patients having the same or
similar angiogenesis-related disorder). The decrease in the rate of
formation of a new blood vessels in a tissue in need thereof in a
patient having an angiogenesis-related disorder can be about 1% to
about 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%,
40%, 35%, 30%, 25%, 20%, 15%, 10%, or 5%; about 5% to about 100%,
95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%,
30%, 25%, 20%, 15%, or 10%; about 10% to about 100%, 95%, 90%, 85%,
80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, or
15%; about 15% to 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%,
55%, 50%, 45%, 40%, 35%, 30%, 25%, or 20%; about 20% to 100%, 95%,
90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, or
25%; about 25% to 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%,
55%, 50%, 45%, 40%, 35%, or 30%; about 30% to 100%, 95%, 90%, 85%,
80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, or 35%; about 35% to
100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, or
40%; about 40% to 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%,
55%, 50%, or 45%; about 45% to 100%, 95%, 90%, 85%, 80%, 75%, 70%,
65%, 60%, 55%, or 50%; about 50% to 100%, 95%, 90%, 85%, 80%, 75%,
70%, 65%, 60%, or 55%; about 55% to 100%, 95%, 90%, 85%, 80%, 75%,
70%, 65%, or 60%; about 60% to 100%, 95%, 90%, 85%, 80%, 75%, 70%,
or 65%; about 65% to 100%, 95%, 90%, 85%, 80%, 75%, or 70%; about
70% to 100%, 95%, 90%, 85%, 80%, or 75%; about 75% to 100%, 95%,
90%, 85%, or 80%; about 80% to 100%, 95%, 90%, or 85%; about 85% to
100%, 95%, or 90%; about 90% to about 100% or 95%; or about 95% to
about 100% (e.g., as compared to the rate of formation of new blood
vessels in the tissue in the patient prior to treatment, or the
rate of formation of new blood vessels in a patient or a population
of patients having the same or similar angiogenesis-related
disorder).
[0325] Also provided is the use of a compound of General Formula I
or a pharmaceutically acceptable salt or solvent thereof for
treating an angiogenesis-related disorder in a patient. Also
provided is the use of a compound of General Formula I or a
pharmaceutically acceptable salt or solvate thereof in the
manufacture of a medicament for treating an angiogenesis-related
disorder in a patient.
[0326] Also provided are methods for treating cancer in a patient
in need thereof, the method comprising: (a) determining if the
cancer in the patient is an FGFR-associated cancer (e.g., using a
regulatory-agency approved, e.g., FDA-approved, kit for identifying
dysregulation of dysregulation of a FGFR gene, a FGFR protein, or
expression or activity or level of any of the same, in a patient or
a biopsy sample from the patient, or by performing any of the
non-limiting examples of assays described herein); and (b) if the
cancer in the patient is determined to be an FGFR-associated
cancer, administering to the patient a therapeutically effective
amount of a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof.
[0327] Also provided is use of a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof for use in
treating an FGFR-associated cancer (e.g., any of the
FGFR-associated cancers described herein) in a patient identified
or diagnosed as having an FGFR-associated cancer through a step of
performing an assay (e.g., an in vitro assay) (e.g., an assay that
utilizes next generation sequencing, immunohistochemistry, or break
apart FISH analysis) (e.g., using a regulatory agency-approved,
e.g., FDA-approved kit) on a sample obtained from the patient to
determine whether the patient has dysregulation of a FGFR gene, a
FGFR protein, or expression or activity or level of any of the
same, where the presence of dysregulation of a FGFR gene, a FGFR
protein, or expression or activity or level of any of the same,
identifies that the patient has an FGFR-associated cancer. Also
provided is the use of a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof for the
manufacture of a medicament for treating an FGFR-associated cancer
in a patient identified or diagnosed as having an FGFR-associated
cancer through a step of performing an assay (e.g., an in vitro
assay) (e.g., an assay that utilizes next generation sequencing,
immunohistochemistry, or break apart FISH analysis) (e.g., using a
regulatory agency-approved, e.g., FDA-approved, kit) on a sample
obtained from the patient to determine whether the patient has a
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity or level of any of the same where the presence of
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity or level of any of the same, identifies that the patient
has an FGFR-associated cancer. Some embodiments of any of the
methods or uses described herein further include recording in the
patient's clinical record (e.g., a computer readable medium) that
the patient determined to have dysregulation of a FGFR gene, a FGFR
protein, or expression or activity or level of any of the same,
through the performance of the assay, should be administered a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof.
[0328] Also provided is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof, for use in the
treatment of a cancer (e.g., an FGFR-associated cancer) in a
patient in need thereof or a patient identified or diagnosed as
having an FGFR-associated cancer (e.g., a patient that has been
identified or diagnosed as having an FGFR-associated cancer through
the use of a regulatory agency-approved, e.g., FDA-approved, kit
for identifying dysregulation of a FGFR gene, a FGFR protein, or
expression or activity or level of any of the same, in a patient or
a biopsy sample from the sample) (e.g., any of the FGFR-associated
cancers described herein or known in the art). Also provided is the
use of a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof for the manufacture of a medicament for
treating a cancer (e.g., an FGFR-associated cancer) in a patient
identified or diagnosed as having an FGFR-associated cancer (e.g.,
a patient that has been identified or diagnosed as having an
FGFR-associated cancer through the use of a regulatory
agency-approved, e.g., FDA-approved, kit for identifying
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity or level of any of the same, in a patient or a biopsy
sample from the patient) (e.g., any of the FGFR-associated cancers
described herein or known in the art).
[0329] In some embodiments of any of the methods or uses described
herein, the patient has been identified or diagnosed as having a
cancer with dysregulation of a FGFR gene, a FGFR protein, or
expression or activity or level of any of the same (e.g., as
determined using a regulatory agency-approved, e.g., FDA-approved,
assay or kit). In some embodiments of any of the methods or uses
described herein, the patient has a tumor that is positive for
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity or level of any of the same (e.g., as determined using a
regulatory-agency-approved assay or kit). In some embodiments of
any of the methods or uses described herein, the patient can be a
patient with a tumor(s) that is positive for dysregulation of a
FGFR gene, a FGFR protein, or expression or activity or level of
any of the same (e.g., identified as positive using a regulatory
agency-approved, e.g., FDA-approved, assay or kit). In some
embodiments of any of the methods or uses described herein, the
patient can be a patient whose tumors have dysregulation of a FGFR
gene, a FGFR protein, or expression or activity or level of any of
the same (e.g., where the tumor is identified as such using a
regulatory agency-approved, e.g., FDA-approved, kit or assay). In
some embodiments of any of the methods or uses described herein,
the patient is suspected of having an FGFR-associated cancer. In
some embodiments of any of the methods or uses described herein,
the patient has a clinical record indicating that the patient has a
tumor that has dysregulation of a FGFR gene, a FGFR protein, or
expression or activity or level of any of the same (and optionally
the clinical record indicates that the patient should be treated
with any of the compounds of Formula I or a pharmaceutically
acceptable salts or solvates thereof or compositions provided
herein).
[0330] Also provided herein are methods of selecting a treatment
for a patient that include administration of a therapeutically
effective amount of a compound of Formula I or a pharmaceutically
acceptable salt or solvate thereof that include a step of
performing an assay (e.g., an in vitro assay) (e.g., an assay that
utilizes next generation sequencing, immunohistochemistry, or break
apart FISH analysis) (e.g., using a regulatory agency-approved,
e.g., FDA-approved, kit) on a sample obtained from the patient to
determine whether the patient has dysregulation of a FGFR gene, a
FGFR protein, or expression or activity or level of any of the
same, identifying or diagnosing a patient determined to have
dysregulation of a FGFR gene, a FGFR protein, or expression or
activity or level of any of the same, as having an FGFR-associated
cancer, and selecting a therapeutic treatment including
administration of a therapeutically effective amount of a compound
of Formula I or a pharmaceutically acceptable salt or solvate
thereof to the patient identified or diagnosed as having an
FGFR-associated cancer. Some embodiments further include
administering the selected treatment to the patient identified or
diagnosed as having an FGFR-associated cancer.
[0331] In some embodiments of any of the methods or uses described
herein, the cancer (e.g., FGFR-associated cancer) is a
hematological cancer. In some embodiments of any of the methods or
uses described herein, the cancer (e.g., an FGFR-associated cancer)
is a solid tumor. In some embodiments of any of the methods or uses
described herein, the cancer (e.g., FGFR-associated cancer) is any
of the exemplary cancers (e.g., any of the exemplary
FGFR-associated cancers) described herein.
[0332] Also provided herein is a method of treating a disease or
disorder mediated by FGFR (e.g., dysregulation of a FGFR gene, a
FGFR protein, or expression or activity or level of any of the
same) in a patient in need of such treatment, the method comprising
administering to the patient a therapeutically effective amount of
a compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof or a pharmaceutical composition thereof. For
example, the FGFR-associated disease can be any of the
FGFR-associated cancers described herein or known in the art.
[0333] Although the genetic basis of tumorigenesis may vary between
different cancer types, the cellular and molecular mechanisms
required for metastasis appear to be similar for all solid tumor
types. During a metastatic cascade, the cancer cells lose growth
inhibitory response, undergo alterations in adhesiveness and
produce enzymes that can degrade extracellular matrix components.
This leads to detachment of tumor cells from the original tumor,
infiltration into the circulation through newly formed vasculature
(e.g., lymph vessels or blood vessels), migration and extravasation
of the tumor cells at favorable distant sites, where they may form
colonies. A number of genes have been identified as being promoters
or suppressors of metastasis. FGFR proteins have been implicated
for a role in metastasis (Qian et al., Oncogene 33:3411-3421,
2014).
[0334] Accordingly, also provided herein are methods for
inhibiting, preventing, aiding in the prevention, or decreasing the
symptoms of metastasis of a cancer (e.g., a FGFR-associated cancer)
in a patient in need thereof, the method comprising administering
to the patient a therapeutically effective amount of a compound of
Formula I or a pharmaceutically acceptable salt or solvate thereof.
Such methods can be used in the treatment of one or more of the
cancers described herein. In some embodiments, the cancer is an
FGFR-associated cancer. In some embodiments, the compound of
Formula I or a pharmaceutically acceptable salt or solvate thereof
is used in combination with an additional therapy or another
therapeutic agent, including a chemotherapeutic agent, such as a
kinase inhibitor.
[0335] Also provided is a method for inhibiting activity of FGFR1,
FGFR2, FGFR3 and/or FGFR4 in a mammalian cell, comprising
contacting the cell with a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof. In one
embodiment, the contacting is performed in vitro. In another
embodiment, the contacting is performed in vivo (e.g., in a human).
In one embodiment, when the contacting is performed in vivo, the
method can include administering an effective amount of a compound
of Formula I or a pharmaceutically acceptable salt or solvate
thereof to a subject. In some embodiments, the mammalian cell is a
mammalian cancer cell. In one embodiment, the mammalian cancer cell
is any cancer as described herein. In some embodiments, the
mammalian cancer cell is an FGFR-associated cancer cell.
[0336] As used herein, the term "contacting" refers to the bringing
together of indicated moieties in an in vitro system or an in vivo
system. For example, "contacting" an FGFR with a compound provided
herein includes the administration of a compound provided herein to
an individual or patient, such as a human, having FGFR1, FGFR2,
FGFR3 and/or FGFR4, as well as, for example, introducing a compound
provided herein into a sample containing a cellular or purified
preparation containing the FGFR1, FGFR2, FGFR3 and/or FGFR4.
[0337] In the field of medical oncology it is normal practice to
use a combination of different forms of treatment to treat each
patient with cancer. In medical oncology the other component(s) of
such conjoint treatment or therapy in addition to compositions
provided herein may be, for example, surgery, radiotherapy, and
chemotherapeutic agents, such as kinase inhibitors, signal
transduction inhibitors and/or monoclonal antibodies. Compounds of
Formula I therefore may also be useful as adjuvants to cancer
treatment, that is, they can be used in combination with one or
more additional therapies or therapeutic agents, for example a
chemotherapeutic agent that works by the same or by a different
mechanism of action.
[0338] In some embodiments of any the methods described herein, the
compound of Formula I (or a pharmaceutically acceptable salt or
solvate thereof) is administered in combination with a
therapeutically effective amount of at least one additional
therapeutic agent selected from one or more additional therapies or
therapeutic (e.g., chemotherapeutic) agents. Non-limiting examples
of additional therapeutic agents include: receptor tyrosine
kinase-targeted therapeutic agents, such as afatinib, cabozantinib,
cetuximab, crizotinib, dabrafenib, erlotinib, gefitinib, imatinib,
lapatinib, lestaurtinib, nilotinib, pazopanib, panitumumab,
pertuzumab, sunitinib, AG 879, AZ-23, AZ623, Go 6976, GNF-5837,
GTx-186, GW 441756, LOXO-101, MGCD516, RPI-1, RXDX101, and TSR-011;
FGFR-targeted therapeutic agents, such as signal transduction
pathway inhibitors, such as Ras-Raf-MEK-ERK pathway inhibitors
(e.g., binimetinib, selumetinib, encorafinib, sorafenib,
trametinib, and vemurafenib), PI3K-Akt-mTOR-S6K pathway inhibitors
(e.g. everolimus, rapamycin, perifosine, temsirolimus), other
kinase inhibitors, such as baricitinib, brigatinib, capmatinib,
danusertib, ibrutinib, milciclib, quercetin, regorafenib,
ruxolitinib, semaxanib, AP32788, BLU285, BLU554, INCB39110,
INCB40093, INCB50465, INCB52793, INCB54828, MGCD265, NMS-088,
NMS-1286937, PF 477736, PLX3397, PLX7486, PLX8394, PLX9486,
PRN1008, PRN1371, RXDX103, RXDX106, RXDX108, and TG101209; FGFR
inhibitors (e.g., ARQ-087, AZD-4547, BGJ398, nintadanib (BIBF
1120), BLU9931, brivanib (BMS-582664), CH5183284, Dovitinib
(TKI258, CHIR258), E-3810, EWMD-2076, JNJ-42756493, lenvatinib
((E7080), LY2874455, Orantinib (TSU-68, SU6668), PD089828,
PD166866, PD173074, Ponatinib (AP-24534), Semaxanib (SU5416),
SSR128129E, SU4984, SU5402, SUN11602), AB1010, BAY 1163877,
Debio-1347, FGF401, FIIN-2, HMPL-453, MK-2461, pazopanib (Votrient,
GW-786034), PD161570, PD173074, PF-477736, PHA-739358 (danusertib),
PRN1371, regorafenib (Stivarga), SPP86, and Tyrphostin AG 1296, and
TAS120; checkpoint inhibitors, such as ipilimumab, tremelimumab,
nivolumab, pidilizumab, MPDL3208A, MEDI4736, MSB0010718C,
BMS-936559, BMS-956559, BMS-935559 (MDX-1105), AMP-224, and
pembrolizumab; modulators of the apoptosis pathway (e.g.
obataclax); cytotoxic chemotherapeutics, such as arsenic trioxide,
bleomycin, cabazitaxel, capecitabine, carboplatin, cisplatin,
cyclophosphamide, cytarabine, dacarbazine, daunorubicin, docetaxel,
doxorubicin, etoposide, fluorouracil, gemcitabine, irinotecan,
lomustine, methotrexate, mitomycin C, oxaliplatin, paclitaxel,
pemetrexed, temozolomide, and vincristine; angiogenesis-targeted
therapies, such as aflibercept and bevacizumab; immune-targeted
agents, such as aldesleukin, interferon alfa-2b, ipilimumab,
lambrolizumab, nivolumab, prednisone, sipuleucel-T; radiotherapy,
such as radioiodide therapy, external-beam radiation, and radium
223 therapy.
[0339] Yet other therapeutic agents that can be administered with a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, include FGFR inhibitors such as those described,
for example, LY2874455 (Lilly), dovitinib (TKI258) (Novartis),
BGJ398 (Novartis), AZD4547 (AstraZeneca), ponatinib (Ariad), E-3810
(EOS), JNJ-42756493 (Astex/Janssen), and ARQ 087 (ArQule).
[0340] In some embodiments, the amount of a compound of Formula I
or a pharmaceutically acceptable salt or solvate thereof is, in
combination with the at least one additional therapeutic agent,
effective in treating the cancer (e.g., an FGFR-associated cancer).
The at least one additional therapeutic agent may be administered
with a compound of Formula I or a pharmaceutically acceptable salt
or solvate thereof as part of the same or separate dosage forms,
via the same or different routes of administration, and on the same
or different administration schedules according to standard
pharmaceutical practice known to one skilled in the art.
[0341] Also provided herein is (i) a pharmaceutical combination for
treating cancer in a patient in need thereof, which comprises (a) a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, (b) at least one additional therapeutic agent
(e.g., any of the exemplary additional therapeutic agents described
herein or known in the art), and (c) optionally at least one
pharmaceutically acceptable carrier for simultaneous, separate or
sequential use for the treatment of cancer, wherein the amounts of
the compound of Formula I or pharmaceutically acceptable salt or
solvate thereof and of the additional therapeutic agent are
together effective in treating the cancer; (ii) a pharmaceutical
composition comprising such a combination; (iii) the use of such a
combination for the preparation of a medicament for the treatment
of cancer; and (iv) a commercial package or product comprising such
a combination as a combined preparation for simultaneous, separate
or sequential use; and to a method of treatment of cancer a patient
in need thereof. In one embodiment the patient is a human.
[0342] The term "pharmaceutical combination", as used herein,
refers to a pharmaceutical therapy resulting from the mixing or
combining of more than one active ingredient and includes both
fixed and non-fixed combinations of the active ingredients. The
term "fixed combination" means that a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof and at least
one additional therapeutic agent (e.g., a chemotherapeutic agent),
are both administered to a patient simultaneously in the form of a
single composition or dosage. The term "non-fixed combination"
means that a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof and at least one additional therapeutic
agent (e.g., chemotherapeutic agent) are administered to a patient
as separate compositions or dosages, either simultaneously,
concurrently or sequentially with variable intervening time limits,
wherein such administration provides effective levels of the two or
more compounds in the body of the patient. These also apply to
cocktail therapies, e.g. the administration of three or more active
ingredients
[0343] Accordingly, also provided herein is a method of treating
cancer, comprising administering to a patient in need thereof a
pharmaceutical combination for treating cancer which comprises (a)
a compound of Formula I or pharmaceutically acceptable salt or
solvate thereof, (b) an additional therapeutic agent, and (c)
optionally at least one pharmaceutically acceptable carrier for
simultaneous, separate or sequential use for the treatment of
cancer, wherein the amounts of the compound of Formula I or
pharmaceutically acceptable salt or solvate thereof and the
additional therapeutic agent are together effective in treating the
cancer. In one embodiment, the compound of Formula I or
pharmaceutically acceptable salt or solvate thereof, and the
additional therapeutic agent are administered simultaneously as
separate dosages. In one embodiment, the compound of Formula I or
pharmaceutically acceptable salt or solvate thereof, and the
additional therapeutic agent are administered as separate dosages
sequentially in any order, in jointly therapeutically effective
amounts, e.g. in daily or intermittently dosages. In one
embodiment, compound of Formula I or pharmaceutically acceptable
salt or solvate thereof, and the additional therapeutic agent are
administered simultaneously as a combined dosage.
[0344] Also provided herein is a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof, for use in the
treatment of an FGFR-associated disease or disorder as defined
hereinabove.
[0345] Also provided herein are methods of treating a
FGFR-associated disease (e.g., a FGFR-associated cancer, e.g., any
of the FGFR-associated cancers described herein or known in the
art) in a patient that include: (a) administering to a patient
identified or diagnosed as having an FGFR-associated disease (e.g.,
an FGFR-associated cancer) one or more doses of a first FGFR
inhibitor over a treatment period; (b) determining a level of
phosphate in a biological sample including blood, serum, or plasma
obtained from the patient after the treatment period; (c) selecting
a patient having an elevated level of phosphate in the biological
sample as compared to a reference level of phosphate; and (d)
ceasing administration of the first FGFR inhibitor (or instructing
the selected patient to cease administration) and initiating
administration of a therapeutically effective amount of a compound
of Formula I or pharmaceutically acceptable salt or solvate
thereof, or a pharmaceutical agent or composition comprising a
compound of Formula I or pharmaceutically acceptable salt or
solvate thereof (e.g., any of the pharmaceutical agents or
compositions described herein), to the selected patient. Some
embodiments of these methods can further include identifying or
diagnosing a patient as having a FGFR-associated disease (e.g., a
FGFR-associated cancer) using any of the methods described
herein.
[0346] In certain embodiments of these methods, the treatment
period can be from about 1 day to about 30 days (e.g., from about 1
day to about 15 days; e.g. about 7 days; e.g., from about 16 days
to about 30 days, e.g., about 21 days). In other embodiments of
these methods, the treatment period can be from 30 days to about 12
months (e.g., from about 30 days to about 9 months, from about 30
days to about 6 months, from about 30 days to about 120 days, from
about 30 days to about 90 days, from about 30 days to about 60
days). In still other embodiments, the treatment period is 7 days
or more or 21 days or more (e.g., more than 7 days or more than 21
days to about 12 months, more than 7 days or more than 21 days to
about 9 months, more than 7 days or more than 21 days to about 6
months, more than 7 days or more than 21 days to about 120 days,
more than 7 days or more than 21 days to about 90 days, more than 7
days or more than 21 days to about 60 days, more than 7 days or
more than 21 days to about 30 days).
[0347] In some embodiments of these methods, the treatment period
is at least or about 1 day, at least or about 2 days, at least or
about 3 days, at least or about 4 days, at least or about 5 days,
at least or about 6 days, at least or about 7 days, at least or
about 8 days, at least or about 9 days, at least or about 10 days,
at least or about 11 days, at least or about 12 days, at least or
about 13 days, at least or about 14 days, at least or about 15
days, at least or about 16 days, at least or about 17 days, at
least or about 18 days, at least or about 19 days, at least or
about 20 days, at least or about 21 days, at least or about 22
days, at least or about 23 days, at least or about 24 days, at
least or about 25 days, at least or about 26 days, at least or
about 27 days, at least or about 28 days, at least or about 29
days, at least or about 30 days, at least or about 31 days, at
least or about 45 days, at least or about 60 days, at least or
about 90 days, at least or about 120 days, at least or about 6
months, at least or about 9 months, at least or about 12
months.
[0348] As used herein, the term "first FGFR inhibitor" means an
FGFR inhibitor that is not a compound of Formula I or a salt or
solvate thereof, or a pharmaceutical composition that includes a
compound of Formula I or a salt or solvate thereof. Non-limiting
examples of first FGFR inhibitor include JNJ-42756493 or
BGJ398.
[0349] In some embodiments, the treatment period is at least 7 days
(e.g., at least or about 8 days, at least or about 9 days, at least
or about 10 days, at least or about 11 days, at least or about 12
days, at least or about 13 days, at least or about 14 days, at
least or about 15 days, at least or about 16 days, at least or
about 17 days, at least or about 18 days, at least or about 19
days, at least or about 20 days, at least or about 21 days, at
least or about 22 days, at least or about 23 days, at least or
about 24 days, at least or about 25 days, at least or about 26
days, at least or about 27 days, at least or about 28 days, at
least or about 29 days, or at least or about 30 days), the FGFR
inhibitor is JNJ-42756493, and a daily dose of about 6 mg to about
12 mg (e.g., about 6 mg to about 11 mg, about 10 mg, about 9 mg,
about 8 mg, or about 7 mg; about 7 mg to about 12 mg, about 11 mg,
about 10 mg, about 9 mg, or about 8 mg; about 8 mg to about 12 mg,
about 11 mg, about 10 mg, or about 9 mg; about 9 mg to about 12 mg,
about 11 mg, or about 10 mg; about 10 mg to about 12 mg or about 11
mg; or about 11 mg to about 12 mg) of the first FGFR inhibitor is
administered to the patient over the treatment period.
[0350] In some embodiments, the treatment period is at least 21
days (e.g., at least or about 22 days, at least or about 23 days,
at least or about 24 days, at least or about 25 days, at least or
about 26 days, at least or about 27 days, at least or about 28
days, at least or about 29 days, at least or about 30 days, at
least or about 31 days, at least or about 32 days, at least or
about 33 days, at least or about 34 days, at least or about 35
days, at least or about 36 days, at least or about 37 days, at
least or about 38 days, at least or about 39 days, or at least or
about 40 days) the first FGFR is BGJ398, and a daily dose of about
50 mg to about 125 mg (e.g., about 50 mg to about 120 mg, about 115
mg, about 110 mg, about 105 mg, about 100 mg, about 95 mg, about 90
mg, about 85 mg, about 80 mg, about 75 mg, about 70 mg, about 65
mg, about 60 mg, or about 55 mg; about 55 mg to about 120 mg, about
115 mg, about 110 mg, about 105 mg, about 100 mg, about 95 mg,
about 90 mg, about 85 mg, about 80 mg, about 75 mg, about 70 mg,
about 65 mg, or about 60 mg; about 60 mg to about 120 mg, about 115
mg, about 110 mg, about 105 mg, about 100 mg, about 95 mg, about 90
mg, about 85 mg, about 80 mg, about 75 mg, about 70 mg, or about 65
mg; about 65 mg to about 120 mg, about 115 mg, about 110 mg, about
105 mg, about 100 mg, about 95 mg, about 90 mg, about 85 mg, about
80 mg, about 75 mg, or about 70 mg; about 70 mg to about 120 mg,
about 115 mg, about 110 mg, about 105 mg, about 100 mg, about 95
mg, about 90 mg, about 85 mg, about 80 mg, or about 75 mg; about 75
mg to about 120 mg, about 115 mg, about 110 mg, about 105 mg, about
100 mg, about 95 mg, about 90 mg, about 85 mg, or about 80 mg;
about 80 mg to about 120 mg, about 115 mg, about 110 mg, about 105
mg, about 100 mg, about 95 mg, about 90 mg, or about 85 mg; about
85 mg to about 120 mg, about 115 mg, about 110 mg, about 105 mg,
about 100 mg, about 95 mg, or about 90 mg; about 90 mg to about 120
mg, about 115 mg, about 110 mg, about 105 mg, about 100 mg, or
about 95 mg; about 95 mg to about 120 mg, about 115 mg, about 110
mg, about 105 mg, or about 100 mg; about 100 mg to about 120 mg,
about 115 mg, about 110 mg, or about 105 mg; about 105 mg to about
120 mg, about 115 mg, or about 110 mg; about 110 mg to about 120 mg
or about 115 mg; or about 115 mg to about 120 mg) of the first FGFR
inhibitor is administered to the patient over the treatment
period.
[0351] Hyperphosphatemia refers to an abnormally elevated level of
phosphate in the blood. In some embodiments, the presence of
hyperphosphatemia in a subject (e.g., a patient) can be determined
by measuring a level(s) of phosphate in a biological sample
including blood, serum, or plasma (e.g., peripheral blood) obtained
from the patient after a particular treatment period (e.g., any of
the treatment periods described herein). Determining the phosphate
level in peripheral blood can be achieved using conventional
methods known in the art (see, e.g., serum phosphate test offered,
e.g., by the Mayo Clinic Laboratories, which utilizes the Roche
Phosphorus reagent (Roche Diagnostics, Inc.; the test is based on
the reaction of phosphate with ammonium molybdate to form ammonium
phosphomolybdate (without reduction)).
[0352] In certain embodiments, the serum phosphate level exhibited
by a subject (e.g., a subject treated with a first FGFR inhibitor;
e.g., a subject selected in step (c) above) is at least or about 5
mg/dL, at least or about 5.5 mg/dL, at least or about 6.0 mg/dL, at
least or about 6.5 mg/dL, at least or about 7.0 mg/dL, at least or
about 7.5 mg/dL, at least or about 8.0 mg/dL, at least or about 8.5
mg/dL, at least or about 9.0 mg/dL, at least or about 9.5 mg/dL, at
least or about 10 mg/dL, at least or about 10.5 mg/dL, at least or
about 11 mg/dL, at least or about 11.5 mg/dL, at least or about 12
mg/dL, at least or about 12.5 mg/dL, at least or about 13 mg/dL, at
least or about 13.5 mg/dL, at least or about 14 mg/dL, or at least
or about 15 mg/dL. In some embodiments, the reference level of
phosphate can be the level in a healthy subject or the average
level in a population of healthy subjects (e.g., subjects not
having hyperphosphatemia or a subjects not at risk for developing
hyperphosphatemia, such as those having a serum phosphate level of
from about 2.0 mg/dL to about 5.0 mg/dL; e.g., from about 2.5 mg/dL
to about 4.5 mg/dL).
[0353] In some examples, the step (c) further includes selecting a
patient having an elevated level of phosphate in the biological
sample as compared to a reference level of phosphate (e.g., any of
the reference level of phosphate described herein) and one or both
of: (i) a calcium-phosphate product (serum calcium in mg/dL x serum
phosphate in mg/dL) of at least or about 50 mg.sup.2/dL.sup.2
(e.g., at least or about 52 mg.sup.2/dL.sup.2, at least or about 54
mg.sup.2/dL.sup.2, at least or about 56 mg.sup.2/dL.sup.2, at least
or about 58 mg.sup.2/dL.sup.2, at least or about 60
mg.sup.2/dL.sup.2, at least or about 62 mg.sup.2/dL.sup.2, at least
or about 64 mg.sup.2/dL.sup.2, at least or about 66
mg.sup.2/dL.sup.2, at least or about 68 mg.sup.2/dL.sup.2, at least
or about 70 mg.sup.2/dL.sup.2, at least or about 72
mg.sup.2/dL.sup.2, at least or about 74 mg.sup.2/dL.sup.2, at least
or about 76 mg.sup.2/dL.sup.2, at least or about 78
mg.sup.2/dL.sup.2, at least or about 80 mg.sup.2/dL.sup.2, at least
or about 82 mg.sup.2/dL.sup.2, at least or about 84
mg.sup.2/dL.sup.2, at least or about 86 mg.sup.2/dL.sup.2, at least
or about 88 mg.sup.2/dL.sup.2, at least about 90 mg.sup.2/dL.sup.2,
at least or about 92 mg.sup.2/dL.sup.2, at least or about 94
mg.sup.2/dL.sup.2, at least or about 96 mg.sup.2/dL.sup.2, at least
about 98 mg.sup.2/dL.sup.2, or at least about 100
mg.sup.2/dL.sup.2) in the biological sample and (ii) a serum
creatinine level of grade 1 or greater (e.g., grade 2, grade 3) in
the biological sample. Exemplary assays for determining the calcium
level of a biological sample including blood, serum, or plasma are
commercially available from BioVision Inc. (Milpitas, Calif.) and
Sigma-Aldrich (St. Louis, Mo.). Exemplary assays for determining
the creatinine level in a biological sample including blood, serum,
or plasma are commercially available from BioVision Inc. (Milpitas,
Calif.) and Diazyme (Poway, Calif.). In other embodiments, the
subject exhibits a serum phosphate level of greater than about 7.0
mg/dL (e.g., a serum phosphate level of greater than 7 mg/dL
lasting for more than 7 days despite phosphate-lowering therapies).
In still other embodiments, the subject exhibits a serum phosphate
level of greater than about 9.0 mg/dL (e.g., a serum phosphate
level of greater than about 9.0 mg/dL for any duration despite
phosphate-lowering therapies). In still other embodiments, the
subject exhibits a serum phosphate level of greater than about 10.0
mg/dL (e.g., a serum phosphate level of greater than about 10.0
mg/dL for any duration).
[0354] In some embodiments, the patient is administered a
therapeutically effective amount of a phosphate binder over the
treatment period. Non-limiting examples of phosphate binders
include aluminum salts (e.g., Alucaps and Basaljel), calcium
carbonate (e.g., Calcichew and Titralac), calcium acetate (e.g.,
Lenal Ace and PhosLo), sevelamer hydrochloride (e.g., Renegel or
Renvela), and lanthanum carbonate (e.g., Fosrenol). In some
embodiments, the patient is administered a therapeutically
effective amount of a phosphate binder each day over the treatment
period. The phosphate binder can be administered at a total daily
dose of about 2.0 g to about 5.0 g (e.g., about 2.0 g to about 4.8
g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g, about 3.8 g,
about 3.6 g, about 3.4 g, about 3.2 g, about 3.0 g, about 2.8 g,
about 2.6 g, about 2.4 g, or about 2.2 g; about 2.2 g to about 5.0
g, about 4.8 g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g,
about 3.8 g, about 3.6 g, about 3.4 g, about 3.2, about 3.0 g,
about 2.8 g, about 2.6 g, or about 2.4 g; about 2.4 to about 5.0 g,
about 4.8 g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g,
about 3.8 g, about 3.6 g, about 3.4 g, about 3.2, about 3.0 g,
about 2.8 g, or about 2.6 g; about 2.6 g to about 5.0 g, about 4.8
g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g, about 3.8 g,
about 3.6 g, about 3.4 g, about 3.2, about 3.0 g, or about 2.8 g;
about 2.8 g to about 5.0 g, about 4.8 g, about 4.6 g, about 4.4 g,
about 4.2 g, about 4.0 g, about 3.8 g, about 3.6 g, about 3.4 g,
about 3.2, or about 3.0 g; about 3.0 g to about 5.0 g, about 4.8 g,
about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g, about 3.8 g,
about 3.6 g, about 3.4 g, or about 3.2 g; about 3.2 g to about 5.0
g, about 4.8 g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g,
about 3.8 g, about 3.6 g, or about 3.4 g; about 3.4 g to about 5.0
g, about 4.8 g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g,
about 3.8 g, or about 3.6 g; about 3.6 g to about 5.0 g, about 4.8
g, about 4.6 g, about 4.4 g, about 4.2 g, about 4.0 g, or about 3.8
g; about 3.8 g to about 5.0 g, about 4.8 g, about 4.6 g, about 4.4
g, about 4.2 g, or about 4.0 g; about 4.0 g to about 5.0 g, about
4.8 g, about 4.6 g, about 4.4 g, or about 4.2 g; about 4.2 g to
about 5.0 g, about 4.8 g, about 4.6 g, or about 4.4 g; about 4.4 g
to about 5.0 g, about 4.8 g, or about 4.6 g; about 4.6 g to about
5.0 g or about 4.8 g; or about 4.8 g to about 5.0 g) over the
treatment period. In some embodiments of these methods, step (d)
further includes ceasing administration of the phosphate binder to
the selected patient or instructing the selected patient to cease
administration of the phosphate binder. In some embodiments of
these methods, step (d) further includes administering a decreased
dose of the phosphate binder to the selected patient relative to
the dose of the phosphate binder administered to the patient over
the treatment period.
[0355] Also provided herein are methods of treating a
FGFR-associated cancer (e.g., any of the FGFR-associated cancers
described herein or known in the art) in a patient that includes
administering a therapeutically effective dose of a compound of
Formula I or a pharmaceutically acceptable salt or solvate thereof,
or a pharmaceutical composition including a compound of Formula I
or a pharmaceutically acceptable salt or solvate thereof, to a
patient identified or diagnosed as having an FGFR-associated cancer
over a treatment period of at least 8 days, where the patient is
determined to have about the same or a decreased level of phosphate
in one or more biological sample(s) including blood, serum, or
plasma obtained from the patient over the treatment period as
compared to a reference level of phosphate (e.g., any of the
reference levels of phosphate described herein). In some
embodiments of any of these methods, the patient is identified or
diagnosed as having a FGFR-associated cancer using any of the
methods described herein or known in the art. Some embodiments of
any of these methods can further include identifying or diagnosing
a subject as having a FGFR-associated cancer using any of the
methods described herein or known in the art. In some embodiments,
the treatment period of at least 8 days can be any of the exemplary
treatment periods (or ranges of treatment periods) described
herein. In some embodiments, the patient is administered a daily
dose of a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof, or or a pharmaceutical composition
including a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof (e.g., any of the pharmaceutical
compositions described herein) over the treatment period.
[0356] In some embodiments of these methods, the patient is
administered a low dose of a phosphate binder (e.g., any of the
exemplary phosphate binders described herein or known in the art)
over the treatment period. In some embodiments of these methods,
the phosphate binder is sevelamer hydrochloride. In some
embodiments of these methods, the lose dose of the phosphate binder
(e.g., sevelamer hydrochloride) can be a total daily administration
of about 0.1 g to about 2.0 g (e.g., about 0.1 g to about 1.9 g,
about 1.8 g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4 g,
about 1.3 g, about 1.2 g, about 1.1 g, about 1.0 g, about 0.9 g,
about 0.8 g, about 0.7 g, about 0.6 g, about 0.5 g, about 0.4 g,
about 0.3 g, or about 0.2 g; about 0.2 g to about 1.9 g, about 1.8
g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4 g, about 1.3
g, about 1.2 g, about 1.1 g, about 1.0 g, about 0.9 g, about 0.8 g,
about 0.7 g, about 0.6 g, about 0.5 g, about 0.4 g, or about 0.3 g;
about 0.3 g to about 1.9 g, about 1.8 g., about 1.7 g, about 1.6 g,
about 1.5 g, about 1.4 g, about 1.3 g, about 1.2 g, about 1.1 g,
about 1.0 g, about 0.9 g, about 0.8 g, about 0.7 g, about 0.6 g,
about 0.5 g, or about 0.4 g; about 0.4 g to about 1.9 g, about 1.8
g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4 g, about 1.3
g, about 1.2 g, about 1.1 g, about 1.0 g, about 0.9 g, about 0.8 g,
about 0.7 g, about 0.6 g, or about 0.5 g; about 0.5 g to about 1.9
g, about 1.8 g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4
g, about 1.3 g, about 1.2 g, about 1.1 g, about 1.0 g, about 0.9 g,
about 0.8 g, about 0.7 g, or about 0.6 g; about 0.6 g to about 1.9
g, about 1.8 g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4
g, about 1.3 g, about 1.2 g, about 1.1 g, about 1.0 g, about 0.9 g,
about 0.8 g, or about 0.7 g; about 0.7 g to about 1.9 g, about 1.8
g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4 g, about 1.3
g, about 1.2 g, about 1.1 g, about 1.0 g, about 0.9 g, or about 0.8
g; about 0.8 g to about 1.9 g, about 1.8 g., about 1.7 g, about 1.6
g, about 1.5 g, about 1.4 g, about 1.3 g, about 1.2 g, about 1.1 g,
about 1.0 g, or about 0.9 g; about 0.9 g to about 1.9 g, about 1.8
g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4 g, about 1.3
g, about 1.2 g, about 1.1 g, or about 1.0 g; about 1.0 g to about
1.9 g, about 1.8 g., about 1.7 g, about 1.6 g, about 1.5 g, about
1.4 g, about 1.3 g, about 1.2 g, or about 1.1 g; about 1.1 g to
about 1.9 g, about 1.8 g., about 1.7 g, about 1.6 g, about 1.5 g,
about 1.4 g, about 1.3 g, or about 1.2 g; about 1.2 g to about 1.9
g, about 1.8 g., about 1.7 g, about 1.6 g, about 1.5 g, about 1.4
g, or about 1.3 g; about 1.3 g to about 1.9 g, about 1.8 g., about
1.7 g, about 1.6 g, about 1.5 g, or about 1.4 g; about 1.4 g to
about 1.9 g, about 1.8 g., about 1.7 g, about 1.6 g, or about 1.5
g; about 1.5 g to about 1.9 g, about 1.8 g., about 1.7 g, or about
1.6 g; about 1.6 g to about 1.9 g, about 1.8 g., or about 1.7 g;
about 1.7 g to about 2.0 g, about 1.9 g, or about 1.8 g; about 1.8
g to about 2.0 g or about 1.9 g; or about 1.9 g to about 2.0 g) of
the phosphate binder.
[0357] In some embodiments, the patient is determined to have about
the same or a decreased level of phosphate in one or more (e.g.,
two, three, four, five, or six) biological sample(s) including
blood, serum, or plasma obtained from the patient at 1 day, 2 days,
3 days, 4 days, 5 days, 6 days, 7 days (1 week), 8 days, 9 days, 10
days, 11 days, 12 days, 13 days, 14 days (2 weeks), 15 days, 16
days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23
days, 24 days, 25 days, 26 days, 27 days, 28 days, 29 days, 30
days, 31 days, 32 days, 33 days, 34 days, 35 days, 36 days, 37
days, 38 days, 39 days, 40 days, 41 days, 42 days, 43 days, 44
days, 45 days, 46 days, 47 days, 48 days, 49 days, 50 days, 51
days, 52 days, 53 days, 54 days, 55 days, 56 days, 57 days, 58
days, 59 days, 60 days, 61 days, 62 days, 63 days, 64 days, 65
days, 66 days, 67 days, 68 days, 69 days, 70 days, 71 days, 72
days, 73 days, 74 days, 75 days, 76 days, 77 days, 78 days, 79
days, 80 days, 81 days, 82 days, 83 days, 84 days, 85 days, 86
days, 87 days, 88 days, 89 days, 90 days, 91 days, 92 days, 93
days, 94 days, 95 days, 96 days, 97 days, 98 days, 99 days, or 100
days following the start of the treatment period as compared to a
reference level of phosphate (e.g., any of the reference levels of
phosphate described herein).
[0358] Also provided are methods of treating a FGFR-associated
cancer (e.g., any of the FGFR-associated cancers described herein
or known in the art) that include administering a therapeutically
effective dose of a compound of Formula I or a pharmaceutically
acceptable salt or solvate thereof, or a pharmaceutical composition
including a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof (e.g., any of the pharmaceutical
compositions described herein) to a patient identified or diagnosed
as having an FGFR-associated cancer over a treatment period (e.g.,
any of the treatment periods described herein), wherein the patient
is not administered a phosphate binder (e.g., any of the phosphate
binders described herein or known in the art) over or during the
treatment period. In some embodiments of any of these methods, the
patient is identified or diagnosed as having a FGFR-associated
cancer using any of the methods described herein or known in the
art. Some embodiments of any of these methods can further include
identifying or diagnosing a subject as having a FGFR-associated
cancer using any of the methods described herein or known in the
art. In some embodiments, the treatment period can be any of the
exemplary treatment periods described herein or any of the
exemplary ranges of treatment periods described herein. In some
embodiments, the patient is administered a daily dose of a compound
of Formula I or a pharmaceutically acceptable salt or solvate
thereof, or a pharmaceutical composition including a compound of
Formula I or a pharmaceutically acceptable salt or solvate thereof
(e.g., any of the pharmaceutical compositions described herein)
over the treatment period.
[0359] Also provided herein are methods of treating a
FGFR-associated cancer (e.g., any FGFR-associated cancer described
herein or known in the art) in a patient that include administering
a therapeutically effective dose of a compound of Formula I or
pharmaceutically acceptable salt or solvate thereof, or a
pharmaceutical composition including a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof (e.g., any of
the pharmaceutical compositions described herein) to a patient
identified or diagnosed as having an FGFR-associated cancer over a
treatment period (e.g., any of the treatment periods described
herein), wherein the patient is further administered a low dose of
a phosphate binder (e.g., any of the phosphate binders described
herein, e.g., sevelamer hydrochloride) (e.g., any of the low doses
of a phosphate binder described herein) over or over at least a
part of the treatment period. Some embodiments of any of these
methods can further include identifying or diagnosing a subject as
having a FGFR-associated cancer using any of the methods described
herein or known in the art. In some embodiments, the treatment
period can be any of the exemplary treatment periods described
herein or any of the exemplary ranges of treatment periods
described herein. In some embodiments, the patient is administered
a daily dose of a compound of Formula I or a pharmaceutically
acceptable salt or solvate thereof, or a pharmaceutical composition
including a compound of Formula I or a pharmaceutically acceptable
salt or solvate thereof (e.g., any of the pharmaceutical
compositions described herein) over the treatment period.
[0360] Also provided are methods of treating a patient having a
FGFR-associated cancer (e.g., any of the FGFR-associated cancers
described herein or known in the art) that include administering a
therapeutically effective dose of a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof, or a
pharmaceutical composition including a compound of Formula I of a
pharmaceutically acceptable salt or solvate thereof (e.g., any of
the pharmaceutical compositions described herein) to a patient
identified or diagnosed as having an FGFR-associated cancer over a
treatment period (e.g., any of the treatment periods described
herein), where the patient does not experience or is less likely to
experience one or more (e.g., two, three, four, five, six, seven,
eight, or nine) of soft tissue calcification, stomatitis, dry
mouth, nail changes, fatigue, asthenia, anorexia, malaise, and
muscle aches over the treatment period or after the treatment
period (e.g., as compared to a patient or a population of patients
having the same FGFR-associated cancer and administered a
therapeutically effective dose of a FGFR inhibitor that is not a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, or a pharmaceutical composition including a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, over the same treatment period). Some embodiments
of any of these methods can further include identifying or
diagnosing a subject as having a FGFR-associated cancer using any
of the methods described herein or known in the art. In some
embodiments, the treatment period can be any of the exemplary
treatment periods described herein or any of the exemplary ranges
of treatment periods described herein. In some embodiments, the
patient is administered a daily dose of a compound of Formula I or
a pharmaceutically acceptable salt or solvate thereof, or a
pharmaceutical composition including a compound of Formula I or a
pharmaceutically acceptable salt or solvate thereof (e.g., any of
the pharmaceutical compositions described herein) over the
treatment period.
[0361] In some embodiments of these methods, the patient is not
administered a phosphate binder (e.g., any of the phosphate binders
described herein or known in the art) during the treatment period.
In such methods, the patient can be, e.g., less likely to
experience one or more (e.g., two, three, four, five, six, seven,
eight, or nine) of soft tissue calcification, stomatitis, dry
mouth, nail changes, fatigue, asthenia, anorexia, malaise, and
muscle aches over the treatment period or after the treatment
period (e.g., as compared to a patient or a population of patients
having the same FGFR-associated cancer and administered a
therapeutically effective dose of a FGFR inhibitor that is not a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, or a pharmaceutical composition including a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, and is not administered a phosphate binder, over
the same treatment period).
[0362] In some embodiments of these methods, the patient is
administered a low dose of a phosphate binder (e.g., any of the
phosphate binders described herein, e.g., sevelamer hydrochloride)
(e.g., any of the exemplary low doses of a phosphate binder
described herein). In such methods, the patient can be, e.g., less
likely to experience one or more (e.g., two, three, four, five,
six, seven, eight, or nine) of soft tissue calcification,
stomatitis, dry mouth, nail changes, fatigue, asthenia, anorexia,
malaise, and muscle aches over the treatment period or after the
treatment period (e.g., as compared to a patient or a population of
patients having the same FGFR-associated cancer and administered a
therapeutically effective dose of a FGFR inhibitor that is not a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, or a pharmaceutical composition including a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, and is administered the same low dose of phosphate
binder, over the same treatment period).
[0363] The level of soft tissue calcification can be
detected/determined in a patient by a medical professional using,
e.g., ultrasound, radiography, computed tomography, and magnetic
resonance imaging. The level of stomatitis, dry mouth, nail
changes, fatigue, asthenia, anorexia, malaise, and muscle aches in
a patient can be determined by a medical professional through the
physical examination of the patient and/or interviewing the patient
(e.g., using a survey).
[0364] In some embodiments, the patient is less likely to
experience one or more (e.g., two, three, four, five, six, seven,
eight, or nine) of soft tissue calcification, stomatitis, dry
mouth, nail changes, fatigue, asthenia, anorexia, malaise, and
muscle aches over the treatment period or after the treatment
period (e.g., as compared to a patient or a population of patients
having the same FGFR-associated cancer and administered a
therapeutically effective dose of a FGFR inhibitor that is not a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, or a pharmaceutical composition including a
compound of Formula I or a pharmaceutically acceptable salt or
solvate thereof, and is administered the same low dose of phosphate
binder, over the same treatment period).
[0365] The phrase "effective amount" means an amount of compound
that, when administered to a patient in need of such treatment, is
sufficient to (i) treat a particular disease, condition, or
disorder mediated by FGFR1, FGFR2 FGFR3 and/or FGFR4, (ii)
attenuate, ameliorate, or eliminate one or more symptoms of the
particular disease, condition, or disorder, or (iii) delay the
onset of one or more symptoms of the particular disease, condition,
or disorder described herein. The amount of a compound of Formula I
that will correspond to such an amount will vary depending upon
factors such as the particular compound, disease condition and its
severity, the identity (e.g., weight) of the patient in need of
treatment, but can nevertheless be routinely determined by one
skilled in the art.
[0366] When employed as pharmaceuticals, the compounds provided
herein can be administered in the form of pharmaceutical
compositions. These compositions can be prepared in a manner well
known in the pharmaceutical art, and can be administered by a
variety of routes, depending upon whether local or systemic
treatment is desired and upon the area to be treated.
Administration may be topical (including transdermal, epidermal,
ophthalmic and to mucous membranes including intranasal, vaginal
and rectal delivery), pulmonary (e.g., by inhalation or
insufflation of powders or aerosols, including by nebulizer;
intratracheal or intranasal), oral or parenteral. Parenteral
administration includes intravenous, intraarterial, subcutaneous,
intraperitoneal intramuscular or injection or infusion; or
intracranial, e.g., intrathecal or intraventricular,
administration. Parenteral administration can be in the form of a
single bolus dose, or may be, for example, by a continuous
perfusion pump. Pharmaceutical compositions and formulations for
topical administration may include transdermal patches, ointments,
lotions, creams, gels, drops, suppositories, sprays, liquids and
powders. Conventional pharmaceutical carriers, aqueous, powder or
oily bases, thickeners and the like may be necessary or
desirable
[0367] Also provided herein pharmaceutical compositions which
contain, as the active ingredient, a compound as provided herein or
a pharmaceutically acceptable salt or solvate thereof, in
combination with one or more pharmaceutically acceptable carriers
(excipients). In some embodiments, the composition is suitable for
topical administration. In making the compositions provided herein,
the active ingredient is typically mixed with an excipient, diluted
by an excipient or enclosed within such a carrier in the form of,
for example, a capsule, sachet, paper, or other container. When the
excipient serves as a diluent, it can be a solid, semi-solid, or
liquid material, which acts as a vehicle, carrier or medium for the
active ingredient. Thus, the compositions can be in the form of
tablets, pills, powders, lozenges, sachets, cachets, elixirs,
suspensions, emulsions, solutions, syrups, aerosols (as a solid or
in a liquid medium), ointments containing, for example, up to 10%
by weight of the active compound, soft and hard gelatin capsules,
suppositories, sterile injectable solutions, and sterile packaged
powders. In one embodiment, the composition is formulated for oral
administration. In one embodiment, the composition is formulated as
a tablet or capsule.
[0368] The compositions comprising a compound provided herein or a
pharmaceutically acceptable salt or solvate thereof can be
formulated in a unit dosage form, each dosage containing from about
5 to about 1,000 mg (1 g), more usually about 100 mg to about 500
mg, of the active ingredient. The term "unit dosage form" refers to
physically discrete units suitable as unitary dosages for human
subjects and other patients, each unit containing a predetermined
quantity of active material (i.e., a compound for Formula I as
provided herein) calculated to produce the desired therapeutic
effect, in association with a suitable pharmaceutical
excipient.
[0369] In some embodiments, the compositions provided herein
contain from about 5 mg to about 50 mg of the active ingredient.
One having ordinary skill in the art will appreciate that this
embodies compounds or compositions containing about 5 mg to about
10 mg, about 10 mg to about 15 mg, about 15 mg to about 20 mg,
about 20 mg to about 25 mg, about 25 mg to about 30 mg, about 30 mg
to about 35 mg, about 35 mg to about 40 mg, about 40 mg to about 45
mg, or about 45 mg to about 50 mg of the active ingredient.
[0370] In some embodiments, the compositions provided herein
contain from about 50 mg to about 500 mg of the active ingredient.
One having ordinary skill in the art will appreciate that this
embodies compounds or compositions containing about 50 mg to about
100 mg, about 100 mg to about 150 mg, about 150 mg to about 200 mg,
about 200 mg to about 250 mg, about 250 mg to about 300 mg, about
350 mg to about 400 mg, or about 450 mg to about 500 mg of the
active ingredient.
[0371] In some embodiments, the compositions provided herein
contain from about 500 mg to about 1,000 mg of the active
ingredient. One having ordinary skill in the art will appreciate
that this embodies compounds or compositions containing about 500
mg to about 550 mg, about 550 mg to about 600 mg, about 600 mg to
about 650 mg, about 650 mg to about 700 mg, about 700 mg to about
750 mg, about 750 mg to about 800 mg, about 800 mg to about 850 mg,
about 850 mg to about 900 mg, about 900 mg to about 950 mg, or
about 950 mg to about 1,000 mg of the active ingredient.
[0372] The active compound may be effective over a wide dosage
range and is generally administered in a pharmaceutically effective
amount. It will be understood, however, that the amount of the
compound actually administered will usually be determined by a
physician, according to the relevant circumstances, including the
condition to be treated, the chosen route of administration, the
actual compound administered, the age, weight, and response of the
individual patient, the severity of the patient's symptoms, and the
like. In some embodiments, the active compound is administered at a
dosage of from about 0.001 mg/Kg to about 500 mg/Kg (e.g., from
about 0.001 mg/Kg to about 200 mg/Kg; from about 0.01 mg/Kg to
about 200 mg/Kg; from about 0.01 mg/Kg to about 150 mg/Kg; from
about 0.01 mg/Kg to about 100 mg/Kg; from about 0.01 mg/Kg to about
50 mg/Kg; from about 0.01 mg/Kg to about 10 mg/Kg; from about 0.01
mg/Kg to about 5 mg/Kg; from about 0.01 mg/Kg to about 1 mg/Kg;
from about 0.01 mg/Kg to about 0.5 mg/Kg; from about 0.01 mg/Kg to
about 0.1 mg/Kg; from about 0.1 mg/Kg to about 200 mg/Kg; from
about 0.1 mg/Kg to about 150 mg/Kg; from about 0.1 mg/Kg to about
100 mg/Kg; from about 0.1 mg/Kg to about 50 mg/Kg; from about 0.1
mg/Kg to about 10 mg/Kg; from about 0.1 mg/Kg to about 5 mg/Kg;
from about 0.1 mg/Kg to about 1 mg/Kg; from about 0.1 mg/Kg to
about 0.5 mg/Kg).
[0373] Provided herein are pharmaceutical kits useful, for example,
in the treatment of FGFR-associated diseases or disorders, such as
cancer, which include one or more containers containing a
pharmaceutical composition comprising a therapeutically effective
amount of a compound provided herein. Such kits can further
include, if desired, one or more of various conventional
pharmaceutical kit components, such as, for example, containers
with one or more pharmaceutically acceptable carriers, additional
containers, etc., as will be readily apparent to those skilled in
the art. Instructions, either as inserts or as labels, indicating
quantities of the components to be administered, guidelines for
administration, and/or guidelines for mixing the components, can
also be included in the kit.
[0374] One skilled in the art will recognize that, both in vivo and
in vitro trials using suitable, known and generally accepted cell
and/or animal models are predictive of the ability of a test
compound to treat or prevent a given disorder.
[0375] One skilled in the art will further recognize that human
clinical trials including first-in-human, dose ranging and efficacy
trials, in healthy patients and/or those suffering from a given
disorder, may be completed according to methods well known in the
clinical and medical arts.
EXAMPLES
[0376] The following examples illustrate the invention.
Synthetic Examples
[0377] Synthesis of Synthetic Intermediates
[0378] Abbreviations
TABLE-US-00002 ACN Acetonitrile bis(pinacolato)diboron
4,4,4',4',5,5,5',5'-octamethyl-2,2'-bi(1,3,2-dioxaborolane) Boc
t-butoxycarbonyl Cu(OAc).sub.2 Copper (II) Acetate DCM
Dichloromethane DIPEA N,N-Diisopropylethylamine DMF
N,N-Dimethylformamide Dppf 1,1'- bis(diphenylphosphanyl) ferrocene
Eq equivalent/equivalents Et.sub.2O Diethyl ether EtOAc Ethyl
Acetate EtOH Ethanol GFF paper or GF/F paper Whatman glass
microfiber filter paper H hour/hours HATU
2-(7-Aza-1H-benzotriazole-1-yl)-1,1,3,3-tetramethyluronium
hexafluorophosphate iPrOH Isopropanol KOAc Potassium Acetate MeOH
Methanol Min minute/minutes MsCl methansulfonyl chloride NBS
N-Bromosuccinimide Pd(OAc).sub.2 Palladium (II) Acetate
Pd(PPh.sub.3).sub.4 Tetrakis(triphenylphosphine)palladium (0)
PdCl.sub.2(dppf).cndot.CH.sub.2Cl.sub.2
1,1'-Bis(diphenylphosphino)ferrocene-palladium(II)dichloride
dichloromethane complex TEA Triethylamine TFA Trifluoroacetic acid
THF tetrahydrofuran TLC thin layer chromatography X-Phos
dicyclohexyl(2',4',6'-triisopropyl-[1,1'-biphenyl]-2-yl)phosphine
[0379] Synthesis of Synthetic Intermediates
[0380] Intermediate S1
##STR00031##
(2R,5R)-tert-butyl
5-methyl-2-(((methylsulfonyl)ox)methyl)morpholine-4-carboxylate
[0381] A cold (0.degree. C.) solution of (2R,5R)-tert-butyl
2-(hydroxymethyl)-5-methylmorpholine-4-carboxylate (275 mg, 1.19
mmol) and DIPEA (312 .mu.L, 1.78 mmol) in DCM (6 mL) was treated
with MsCl (110 .mu.L, 1.43 mmol). The resulting mixture was stirred
overnight at ambient temperature. The mixture was partitioned
between saturated NaHCO.sub.3(aq) (30 mL) and DCM (20 mL), and the
aqueous extracts were washed with additional DCM (2.times.10 mL).
The combined organic extracts were washed with brine (10 mL), dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum affording the title compound (367 mg, 99% yield). This
material was of sufficient purity to be used directly without
further purification.
[0382] The following intermediates shown in Table S1 were prepared
according the method used for the synthesis of Intermediate S1
using the appropriate chiral hydroxymethyl-(morpholine)carboxylate
starting materials. The reaction progression in each was followed
by TLC (50% Hexanes/EtOAc, KMnO.sub.4 stain) and reaction times
were adjusted as necessary.
TABLE-US-00003 TABLE S1 Intermediate Structure Name S2 ##STR00032##
tert-butyl (2S,5R)-5-methyl-2-
(((methylsulfonyl)oxy)methyl)morpholine-4- carboxylate S3
##STR00033## tert-butyl (S)-5,5-dimethyl-2-
(((methylsulfonyl)oxy)methyl)morpholine-4- carboxylate S4
##STR00034## tert-butyl (R)-5,5-dimethyl-2-
(((methylsulfonyl)oxy)methyl)morpholine-4- carboxylate S5
##STR00035## tert-butyl (R)-6-(((methylsulfonyl)oxy)methyl)-
7-oxa-4-azaspiro[2.5]octane-4-carboxylate
[0383] Intermediate S6
##STR00036##
(1-cyanocyclopropyl)methyl methanesulfonate
[0384] A solution of 1-(hydroxymethyl)cyclopropanecarbonitrile
(1.14 g, 11.7 mmol) in DCM (24 mL) was treated with TEA (3.50 mL,
25.8 mmol). The resulting reaction mixture was cooled to 0.degree.
C., treated dropwise with MsCl (1.37 mL, 17.6 mmol) and stirred for
1 h at 0.degree. C. The reaction mixture was stirred at ambient
temperature for an additional 2 h before being diluted with
additional DCM (100 mL) and washed with brine (25 mL). The organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated under vacuum to afford the title compound (2.057
g, 100% yield). This material was of sufficient purity to be used
directly without further purification. 1H NMR (400 MHz, DMSO-d6)
.delta. 4.27 (s, 2H), 3.23 (s, 3H), 1.42-1.38 (m, 2H), 1.21-1.18
(m, 2H).
[0385] Intermediate P1
##STR00037##
tert-butyl
(2R,5R)-5-methyl-2-((4-(4,4,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyr-
azol-1-yl)methyl)morpholine-4-carboxylate
[0386] A mixture of
4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (276
mg, 1.42 mmol), (2R,5R)-tert-butyl
5-methyl-2-(((methylsulfonyl)oxy)methyl)morpholine-4-carboxylate
(Intermediate S1; 367 mg, 1.19 mmol), and Cs.sub.2CO.sub.3(s) (966
mg, 2.97 mmol) was suspended in DMF (5.93 mL) and stirred for 1 day
at ambient temperature. The mixture was partitioned between EtOAc
(100 mL) and H.sub.2O (50 mL). The organic extracts were separated
and then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum. The crude residue was purified by silica
chromatography (0-100% EtOAc/Hexanes) to afford the title compound
(483 mg, 100% yield). MS (apci) m/z=408.2 (M+H).
[0387] The following intermediates shown in Table P1 were prepared
according the method used for the synthesis of Intermediate P1,
tert-butyl
(2R,5R)-5-methyl-2-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-p-
yrazol-1-yl)methyl)morpholine-4-carboxylate, using the appropriate
chiral methanesulfonoxy-methyl-morpholine-4-carboxylate starting
materials (Intermediates S2-S5 from Table S1). All compounds were
purified using a method similar to that used for purifying
Intermediate P1 utilizing the appropriate gradient for the silica
chromatography.
TABLE-US-00004 TABLE P1 MS (apci) Intermediate Structure Name m/z
P2 ##STR00038## tert-butyl (2S,5R)- 5-methyl-2-((4- (4,4,5,5-
tetramethyl-1,3,2- dioxaborolan-2- yl)-1H-pyrazol-1- 408.3 (M + H)
yl)methyl)morpholine- 4-carboxylate P3 ##STR00039## tert-butyl
(S)-5,5- dimethyl-2-((4- (4,4,5,5- tetramethyl-1,3,2-
dioxaborolan-2- yl)-1H-pyrazol-1- yl)methyl)morpholine- 422.2 (M +
H) 4-carboxylate P4 ##STR00040## tert-butyl (R)-5,5-
dimethyl-2-((4- (4,4,5,5- tetramethyl-1,3,2- dioxaborolan-2-
yl)-1H-pyrazol-1- yl)methyl)morpholine- 422.2 (M + H) 4-carboxylate
P5 ##STR00041## tert-butyl (R)-6- ((4-(4,4,5,5- tetramethyl-1,3,2-
dioxaborolan-2- yl)-1H-pyrazol-1- yl)methyl)-7-oxa-4-
azaspiro[2.5]octane- 420.2 (M + H) 4-carboxylate
[0388] Intermediate P6
##STR00042##
1-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)methy-
l)cyclopropane-1-carbonitrile
[0389] A solution of
4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (1.52
g, 7.83 mmol) in DMA (31 mL) was treated with
1-cyanocyclopropyl)methyl methanesulfonate (Intermediate S6; 1.99
g, 11.4 mmol), Cs.sub.2CO.sub.3(s) (3.83 g, 11.8 mmol) and 4A
molecular sieves (250 mg). The resulting suspension was stirred for
16 h at 100.degree. C. After cooling to ambient temperature, the
reaction mixture was filtered, and the solids collected were rinsed
with EtOAc (50 mL). The filtrate was diluted with toluene (150 mL)
and concentrated under vacuum. The resulting crude residue was
azeotroped with toluene (150 mL) several times to remove most of
the DMA and subsequently purified by silica chromatography (5-75%
Hexanes/EtOAc as the gradient eluent) to afford the title compound
(1.38 g, 65% yield). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
8.03 (s, 1H), 7.65 (s, 1H), 4.28 (s, 2H), 1.33-1.23 (m, 16H).
[0390] Intermediate P7
##STR00043##
4,4,4-trifluoro-1-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyr-
azol-1-yl)butan-2-ol
[0391] A mixture of
4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (500
mg, 2.58 mmol), 2-(2,2,2-trifluoroethyl)oxirane (390 mg, 3.09
mmol), and Cs.sub.2CO.sub.3(s) (1.68 g, 5.15 mmol) was suspended in
DMF (2.58 mL) and stirred overnight at 80.degree. C. The mixture
was partitioned between EtOAc (50 mL) and H.sub.2O (25 mL). The
organic extracts were separated then dried over anhydrous
Na.sub.2SO.sub.4(s) filtered and concentrated under vacuum to
afford the title compound (825 mg, 100% yield). MS (apci) m/z=321.1
(M+H). This material was of sufficient purity to be used directly
without further purification.
[0392] Intermediate P8
##STR00044##
2-methyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1--
yl)propanenitrile
Step 1: Preparation of
2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)aceton-
itrile
[0393] A solution of
4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (5.11
g, 26.3 mmol) in DMF (50 mL) was treated with bromoacetonitrile
(2.20 mL, 31.6 mmol) and K.sub.2CO.sub.3(s) (5.46 g, 39.5 mmol).
The resulting suspension was stirred for 24 h at 100.degree. C. The
reaction mixture was cooled to ambient temperature, diluted with
water (100 mL) then extracted with EtOAc (3.times.250 mL). The
combined organic extracts were washed with water (3.times.50 mL)
and brine (50 mL) then dried over anhydrous Na.sub.2SO.sub.4(s).
Following filtration, the organic extracts were concentrated under
vacuum then purified by silica chromatography (10-60% Hexanes/EtOAc
as the gradient eluent) to afford the title compound (2.42 g, 39%
yield). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 8.04 (s, 1H),
7.71 (s, 1H), 5.49 (s, 2H), 1.25 (s, 12H).
Step 2: Preparation of
2-methyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1--
yl)propanenitrile
[0394] A cold (0.degree. C.) solution of
2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)aceton-
itrile (2.42 g, 10.4 mmol) in THF (26 mL) was treated with
iodomethane (1.94 mL, 31.1 mmol) then drop-wise with sodium
bis(trimethylsilyl)amide (22.8 mL, 22.8 mmol). The resulting
mixture was stirred 1 h at 0.degree. C. before quenching with the
addition of saturated NH.sub.4Cl.sub.(aq) (25 mL). At ambient
temperature the reaction mixture was then partitioned between EtOAc
(250 mL) and water (100 mL). The organic extracts were washed again
with water (50 mL) and brine (50 mL), then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
crude residue was purified by silica chromatography (5-50%,
Hexanes/EtOAc as the gradient eluent) to afford the title compound
(1.35 g, 50% yield). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
8.20 (s, 1H), 7.75 (s, 1H), 1.97 (s, 6H), 1.26 (s, 12H).
[0395] Intermediate P9
##STR00045##
2-methyl-1-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1--
yl)propan-2-ol
[0396] A mixture of
4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (2.16
g, 11.1 mmol) and Cs.sub.2CO.sub.3(s) (3.81 g, 11.7 mmol) in
2,2-dimethyloxirane (3 mL, 33.6 mmol) was stirred overnight at
100.degree. C. The reaction mixture was filtered through GFF paper
and the filtrate was concentrated under vacuum to afford the title
compound (2.48 g, 84% yield). This material was of sufficient
purity to be used directly without further purification. .sup.1H
NMR (CDCl3) .delta. 7.81 (s, 1H), 7.69 (s, 1H
[0397] Intermediate Y1
##STR00046##
tert-butyl 4-(5-aminopyrazin-2-yl)-1H-pyrazole-1-carboxylate
[0398] A mixture of 2-amino-5-bromopyrazine (100 mg, 0.575 mmol),
tert-butyl
4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole-1-carboxylate
(338 mg, 1.15 mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (47.3 mg,
0.0575 mmol), K.sub.2CO.sub.3(s) (238 mg, 1.72 mmol) was suspended
in a mixture of dioxane (5.75 mL) and water (1.15 mL). The mixture
was sparged with Ar.sub.(g), then sealed and stirred for 4 h at
90.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with EtOAc, filtered then concentrated under
vacuum. The crude residue was purified by silica chromatography
(70-100% EtOAc in Hexanes as the eluent) to afford the title
compound (84 mg, 56% yield). MS (apci) m/z=162.1 (desBoc M+H).
[0399] The following 4-(5-aminopyrazin-2-yl)-1H-pyrazole
intermediates, shown in Table Y1, were prepared in a manner similar
to the method used for the synthesis of Intermediate Y1, using the
appropriate arylboronate starting materials (commercially available
or synthesized according to Examples provided herein), excess
K.sub.2CO.sub.3(s) (0.1-0.2 equivalents), 0.1-0.2 M in 5:1
dioxane:water and temperatures between 85-90.degree. C. Reaction
progression in each was followed by LCMS and reactions times were
adjusted as necessary. All compounds were purified by silica
chromatography as in Intermediate Y1 utilizing the appropriate
eluent.
TABLE-US-00005 TABLE Y1 MS (apci) Intermediate Structure Name m/z
Y9 ##STR00047## (2R,5R)-tert-butyl 2- ((4-(5-aminopyrazin-
2-yl)-1H-pyrazol-1- yl)methyl)-5- methylmorpholine-4- carboxylate
275.1 [(M-Boc) + H)] Y10 ##STR00048## (2S,5R)-tert-butyl 2-
((4-(5-aminopyrazin- 2-yl)-1H-pyrazol-1- yl)methyl)-5-
methylmorpholine-4- carboxylate 275.1 [(M-Boc) + H)] Y11
##STR00049## tert-butyl (S)-2-((4-(5- aminopyrazin-2-yl)-
1H-pyrazol-1- yl)methyl)-5,5- dimethylmorpholine- 4-carboxylate
389.2 (M + H) Y12 ##STR00050## tert-butyl (R)-2-((4-(5-
aminopyrazin-2-yl)- 1H-pyrazol-1- yl)methyl)-5,5-
dimethylmorpholine- 4-carboxylate 389.2 (M + H) Y13 ##STR00051##
tert-butyl (R)-6-((4-(5- aminopyrazin-2-yl)- 1H-pyrazol-1-
yl)methyl)-7-oxa-4- azaspiro[2.5]octane-4- carboxylate 387.2 (M +
H)
[0400] Intermediate Y2
##STR00052##
5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine
[0401] A solution of 2-amino-5-bromopyrazine (5.7 g, 33 mmol) in
4:1 dioxane:water (300 mL) was treated with
1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole
(7.2 g, 34 mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (1.3 g, 1.6
mmol), K.sub.2CO.sub.3(s) (14 g, 98 mmol). The mixture was sparged
with Ar.sub.(g), then sealed and stirred for 16 h at 100.degree. C.
After cooling to ambient temperature, the reaction mixture was
diluted with 4:1 DCM/iPrOH (500 mL), and the resulting solution was
extracted with water (2.times.100 mL). The organic extracts were
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum. The resulting crude residue was purified by silica
chromatography (1-30% DCM/MeOH as the gradient eluent) to afford
the title compound (63.4 mg, 63% yield). MS (apci) m/z=176.1
(M+H).
[0402] Intermediate Y3
##STR00053##
5-(1-isopropyl-1H-pyrazol-4-yl)pyrazin-2-amine
[0403] A mixture of 2-amino-5-bromopyrazine (0.505 g, 2.90 mmol),
1-isopropyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole
(0.822 g, 3.48 mmol), Pd(PPh.sub.3).sub.4 (0.168 g, 0.145 mmol), 2M
Na.sub.2CO.sub.3(aq) (3.05 mL, 6.09 mmol) in dioxane (9 mL) was
stirred overnight at 90.degree. C. After cooling to ambient
temperature, the reaction mixture was diluted with DCM then
extracted with water and brine. The organic extracts were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered then concentrated
under vacuum. The resulting crude residue was purified by silica
chromatography to afford the title compound (0.415 g, 70% yield).
MS (apci) m/z=204.1 (M+H).
[0404] The following 4-(5-aminopyrazin-2-yl)-1H-pyrazole
intermediates, shown in Table Y3, were prepared according the
method used for the synthesis of Intermediate Y3 using the
appropriate arylboronate starting materials (commercially available
or prepared as described herein). Reaction progression in each was
followed by LCMS and reaction time was adjusted as necessary. All
compounds were purified by silica chromatography according to the
method for the isolation of Intermediate Y3 utilizing the
appropriate eluent.
TABLE-US-00006 TABLE Y3 Intermediate MS (apci) # Structure Name m/z
Y4 ##STR00054## 5-(1-isobutyl-1H- pyrazol-4- yl)pyrazin-2-amine
218.1 (M + H) Y6 ##STR00055## 5-(1- (cyclopropylmethyl)-
1H-pyrazol-4- yl)pyrazin-2-amine 216.1 (M + H) Y7 ##STR00056##
5-(1-cyclobutyl- 1H-pyrazol-4- yl)pyrazin-2-amine 216.1 (M + H) Y8
##STR00057## 5-(1-(2- morpholinoethyl)- 1H-pyrazol-4-
yl)pyrazin-2-amine 275.1 (M + H) Y17 ##STR00058## 1-(4-(5-
aminopyrazin-2- yl)-1H-pyrazol-1- yl)-2- methylpropan-2-ol 234.2 (M
+ H)
[0405] Intermediate Y5
##STR00059##
5-(1-(pentan-3-yl)-1H-pyrazol-4-yl)pyrazin-2-amine
[0406] A mixture of 2-amino-5-bromopyrazine (90.6 mg, 0.521 mmol),
1-(pentan-3-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazo-
le (194 mg, 0.573 mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (42.84
mg, 0.0521 mmol), K.sub.2CO.sub.3(s) (216 mg, 1.56 mmol) was
suspended in a mixture of dioxane (5.21 mL), and water (1.04 mL).
The mixture was sparged with Ar.sub.(g), then sealed and stirred
overnight at 90.degree. C. After cooling to ambient temperature,
the reaction mixture was diluted with DCM and water. The organic
extracts were washed with water and brine then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting crude residue was purified by silica chromatography
(70-100% EtOAc in Hexanes as the eluent) to afford the title
compound (40 mg, 33% yield). MS (apci) m/z=232.1 (M+H).
[0407] Intermediate Y14
##STR00060##
1-((4-(5-aminopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)cyclopropane-1-carboni-
trile
[0408] A solution of
1-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)methy-
l)cyclopropanecarbonitrile (Intermediate P6; 560.0 mg, 2.050 mmol)
in 4:1 dioxane:water (10 mL) was treated with
2-amino-5-bromopyrazine (356.7 mg, 2.050 mmol), Pd(PPh.sub.3).sub.4
(236.9 mg, 0.2050 mmol), K.sub.2CO.sub.3(s) (850.1 mg, 6.151 mmol).
The mixture was sparged with Ar.sub.(g), then sealed and stirred
for 16 h at 100.degree. C. After cooling to ambient temperature,
the reaction mixture was diluted with 4:1 DCM:iPrOH (200 mL) and
the resulting solution was extracted with water (2.times.50 mL).
The organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by silica chromatography (1-30% DCM/MeOH as the
gradient eluent) to afford the title compound (246.6 mg, 51%
yield). MS (apci) m/z=241.1 (M+H).
[0409] Intermediate Y15
##STR00061##
1-(4-(5-aminopyrazin-2-yl)-1H-pyrazol-1-yl)-4,4,4-trifluorobutan-2-ol
[0410] The title compound was prepared (83.2 mg, 23% yield)
according to the method described for Intermediate Y14, using
4,4,4-trifluoro-1-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyr-
azol-1-yl)butan-2-ol (Intermediate P7) in place of
1-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)methy-
l)cyclopropanecarbonitrile (Intermediate P6). MS (apci) m/z=288.0
(M+H).
[0411] Intermediate Y16
##STR00062##
2-(4-(5-aminopyrazin-2-yl)-1H-pyrazol-1-yl)-2-methylpropanenitrile
[0412] A solution of 2-amino-5-bromopyrazine (258.9 mg, 1.488 mmol)
in 4:1 dioxane:water (10 mL) was treated with
1-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)methy-
l)cyclopropanecarbonitrile (Intermediate P8; 560.0 mg, 2.050 mmol),
PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (122.4 mg, 0.1488 mmol),
K.sub.2CO.sub.3(s) (616.9 mg, 4.464 mmol). The mixture was sparged
with Ar.sub.(g), then sealed and stirred for 16 h at 100.degree. C.
After cooling to ambient temperature, the reaction mixture was
diluted with 4:1 DCM:iPrOH (250 mL) and the resulting solution was
extracted with water (2.times.50 mL). The organic extracts were
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum. The resulting crude residue was purified by C18
reverse phase chromatography (5-95% water/ACN with 0.1% TFA as the
gradient eluent) to afford the title compound as a TFA salt. The
TFA salt was dissolved in 4:1 DCM:iPrOH (100 mL) and extracted with
saturated NaHCO.sub.3(aq) (1.times.25 mL). The organic extracts
were dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum to cleanly afford the title compound (77
mg, 23% yield). MS (apci) m/z=229.1 (M+H).
[0413] Intermediate Y18
##STR00063##
2-(4-(5-
aminopyrazin-2-yl)-1H-pyrazol-1-yl)-2-methylpropan-1-ol
[0414] The title compound was prepared (97.2 mg, 23% yield)
according to the method described for Intermediate Y16, using
2-methyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1--
yl)propan-1-ol in place of
1-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)methy-
l)cyclopropanecarbonitrile (Intermediate P9). MS (apci) m/z=234.1
(M+H).
[0415] Intermediate Y19
##STR00064##
tert-butyl
4-(4-(5-aminopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0416] A mixture of 2-amino-5-bromopyrazine (5.0 g, 28.7 mmol),
tert-butyl
4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)piperi-
dine-1-carboxylate (11.9 g, 31.6 mmol), Pd(PPh.sub.3).sub.4 (3.32
g, 2.87 mmol), 2M Na.sub.2CO.sub.3(aq) (35.9 mL, 71.8 mmol) in
dioxane (57.5 mL) was purged with N.sub.2(g) for 6 min then sealed
and stirred for 16 h at 90.degree. C. After cooling to ambient
temperature, the reaction mixture was diluted with EtOAc (300 mL)
and washed with water (2.times.80 mL). The combined organic
extracts were dried over anhydrous MgSO.sub.4(s), filtered and
concentrated under vacuum. The resulting crude residue was
precipitated from hot ACN (X mL) to provide pure title compound
(6.35 g). Mother liquor was concentrated under vacuum and the
residue obtained was purified by flash chromatography on silica gel
(Redi Sep 220 g) eluting with 5-60% acetone/DCM (15CV) to provide
additional title compound (3.17 g; 96% total yield). MS (apci)
m/z=245.1 [(M-Boc)+H]. MS data are for the purified forms of batch
land batch 2.
[0417] Intermediate Y20
##STR00065##
5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-amine
[0418] A solution of 2-amino-5-bromopyrazine 1.03 g, 5.95 mmol) in
4:1 dioxane:water (20 mL) was treated with
1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole
(1.30 g, 6.25 mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (0.490 g,
0.595 mmol), K.sub.2CO.sub.3(s) (2.47 g, 17.9 mmol). The mixture
was sparged with Ar.sub.(g), then sealed and stirred for 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (250 mL) and the resulting
solution was extracted with water (2.times.50 mL). The organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated under vacuum. The resulting crude residue was
purified by silica chromatography (1-30% DCM/MeOH as the gradient
eluent) to afford the title compound (845.5 mg, 82% yield). MS
(apci) m/z=175.1 (M+H).
[0419] Intermediate Y21
##STR00066##
tert-butyl
4-(4-(6-aminopyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0420] The title compound was prepared (1.12 g, 85% yield)
according to the method described for Intermediate Y21, using
tert-butyl
4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)piperi-
dine-1-carboxylate in place of
1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole
and a step wise gradient eluent system of 5-60% DCM/EtOAc then
1-25% DCM/MeOH in the silica chromatography. MS (apci) m/z=344.1
(M+H).
[0421] Intermediate L1
##STR00067##
tert-butyl
4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazole-1-carboxylate
[0422] A solution of tert-butyl
4-(5-aminopyrazin-2-yl)-1H-pyrazole-1-carboxylate (Intermediate Y1;
84 mg, 0.32 mmol) in CHCl.sub.3 (3.2 mL) was treated with pyridine
(29 .mu.L, 0.35 mmol) and the resulting solution was cooled to
0.degree. C. Br.sub.2 (17 .mu.L, 0.34 mmol) was added dropwise to
the solution and the resulting reaction mixture was stirred at
0.degree. C. for 30 min. The reaction mixture was then stirred at
ambient temperature for 2 h prior to quenching with 10%
Na.sub.2S.sub.2O.sub.3(aq). The resulting biphasic mixture was
extracted with DCM. The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting crude residue was purified by silica chromatography
(30-100% EtOAc in Hexanes as the gradient eluent) to afford the
title compound (69 mg, 63% yield). MS (apci) m/z=242.0
[(M-Boc)+H+2], 240 [(M-Boc)+H], with Br pattern.
[0423] The following intermediates shown in Table L1 were prepared
according the method used for the synthesis of Intermediate L1.
Reaction progression in each was followed by LCMS and reaction time
was adjusted as necessary. All compounds were purified by silica
chromatography according the method used for purifying Intermediate
L1 using the appropriate gradient eluent.
TABLE-US-00007 TABLE L1 Intermediate # Structure Name MS (apci) m/z
L5 ##STR00068## 3-bromo-5-(1- (pentan-3-yl)-1H- pyrazol-4-
yl)pyrazin-2-amine 312.0 ([(M + H) + 2] 310 (M + H) (with bromine
pattern) L19 ##STR00069## tert-butyl 4-(4-(5- amino-6-
bromopyrazin-2- yl)-1H-pyrazol-1- yl)piperidine-1- carboxylate
369.0 ([(des(tBu)M + H) + 2] 367 (des(tBu)M + H) (with bromine
pattern)
[0424] Intermediate L2
##STR00070##
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine
[0425] A cold (0.degree. C.) solution of
5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate Y2; 0.210
g, 1.20 mmol) and pyridine (0.107 mL, 1.32 mmol) in CHCl.sub.3 (10
mL) was treated a solution of Br.sub.2 (0.422 g, 2.64 mmol) in
CHCl.sub.3 (4 mL). The resulting reaction mixture was maintained at
0.degree. C. for 5 min and then allowed to stir at ambient
temperature for 2 h. The reaction mixture was then diluted with DCM
(50 mL) prior to quenching with saturated
Na.sub.2S.sub.2O.sub.3(aq) (20 mL). The resulting biphasic mixture
was separated, the organic extracts were reserved and the aqueous
extracts were washed with DCM (50 mL). The combined organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated under vacuum. The resulting crude residue was
purified by silica chromatography (1:1 EtOAc/Hexanes as the eluent)
to afford the title compound (0.266 g, 87% yield). MS (apci)
m/z=256.0 [(M+H)+2], 254.0 (M+H), with Br pattern.
[0426] Intermediate L3
##STR00071##
3-bromo-5-(1-isopropyl-1H-pyrazol-4-yl)pyrazin-2-amine
[0427] A cold (0.degree. C.) solution of
5-(1-isopropyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate Y3;
0.365 g, 1.80 mmol) and pyridine (0.160 mL, 1.98 mmol) in
CHCl.sub.3 (12 mL) was treated with Br.sub.2 (0.0971 mL, 1.89
mmol). The reaction mixture was stirred overnight at ambient
temperature then diluted with DCM and extracted with saturated
Na.sub.2S.sub.2O.sub.3(aq). The organic extracts were washed with
brine, then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum. The resulting crude residue was purified
by silica chromatography to afford the title compound (0.247 g, 49%
yield). MS (apci) m/z=282.0 (M+H), 284.0 [[(M+H)+2]](bromine
pattern).
[0428] The following intermediates shown in Table L3 were prepared
according the method used for the synthesis of Intermediate L3 in
CHCl.sub.3 (0.1-0.15 M). Reaction progression in each was followed
by LCMS and reaction time was adjusted as necessary. All compounds
were purified by silica chromatography according the method used
for isolating Intermediate L3 using the appropriate gradient
eluent.
TABLE-US-00008 TABLE L3 Intermediate # Structure Name MS (apci) m/z
L4 ##STR00072## 3-bromo-5-(1-isobutyl- 1H-pyrazol-4-
yl)pyrazin-2-amine 296.0 (M + H), 298.0 [(M + H) + 2] (with bromine
pattern) L6 ##STR00073## 3-bromo-5-(1- (cyclopropylmethyl)-
1H-pyrazol-4- yl)pyrazin-2-amine 294.0 (M + H), 296.0 [[(M + H) +
2]] (with bromine pattern) L7 ##STR00074## 3-bromo-5-(1-
cyclobutyl-1H-pyrazol- 4-yl)pyrazin-2-amine 296.0 [[(M + H) + 2]],
294.0 (M + H) (with bromine pattern) L8 ##STR00075##
3-bromo-5-(1-(2- morpholinoethyl)-1H- pyrazol-4-yl)pyrazin-2- amine
354.9 [(M + H) + 2], 353.0 (M + H) (with bromine pattern) L17
##STR00076## 1-(4-(5-amino-6- bromopyrazin-2-yl)-1H-
pyrazol-1-yl)-2- methylpropan-2-ol 314.09 [[(M + H) + 2]], 312.0 (M
+ H) (with bromine pattern)
[0429] Intermediate L10
##STR00077##
tert-butyl
(2S,5R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5-met-
hylmorpholine-4-carboxylate
[0430] A solution of (2S,5R)-tert-butyl
2-((4-(5-aminopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5-methylmorpholine-4--
carboxylate (Intermediate Y10; 150 mg, 0.401 mmol) and pyridine
(35.61 .mu.L, 0.441 mmol) in CHCl.sub.3 (4.01 mL) was cooled to
0.degree. C., stirred for 30 min, then treated dropwise with
Br.sub.2 (21.552 .mu.L, 0.421 mmol). The reaction mixture, then was
stirred overnight at ambient temperature prior to quenching with
10% Na.sub.2S.sub.2O.sub.3(aq). The resulting biphasic mixture was
extracted with DCM (3x). The organic extracts were washed with
water (2x) and brine then dried over anhydrous MgSO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by silica chromatography (60-100% EtOAc/Hexanes as the
gradient eluent) to afford the title compound (50 mg, 28% yield).
MS (apci) m/z=455.0 [(M+H)+2], 453 (M+H), with Br pattern.
[0431] The following intermediates shown in Table L10 were prepared
according the method used for the synthesis of Intermediate L10 in
CHCl.sub.3 (0.1-0.2 M), from the appropriate starting materials
prepared as described herein. Reaction progression was followed by
LCMS and reaction time was adjusted as necessary. All compounds
were purified by silica chromatography as in Intermediate L10 using
the appropriate gradient eluent.
TABLE-US-00009 TABLE L10 Intermediate # Structure Name MS (apci)
m/z L9 .sup.a ##STR00078## tert-butyl (2R,5R)-2- ((4-(5-amino-6-
bromopyrazin-2-yl)- 1H-pyrazol-1- yl)methyl)-5- methylmorpholine-4-
carboxylate 455.0 [(M + H) + 2] 453 (M + H) L11 ##STR00079##
tert-butyl (S)-2-((4- (5-amino-6- bromopyrazin-2-yl)- 1H-pyrazol-1-
yl)methyl)-5,5- dimethylmorpholine- 4-carboxylate 467.0 (M+), 469.1
(M + 2) L12 ##STR00080## tert-butyl (R)-2-((4- (5-amino-6-
bromopyrazin-2-yl)- 1H-pyrazol-1- yl)methyl)-5,5-
dimethylmorpholine- 4-carboxylate 467.1 (M+), 469.1 (M + 2) L13
##STR00081## tert-butyl (R)-6-((4- (5-amino-6- bromopyrazin-2-yl)-
1H-pyrazol-1- yl)methyl)-7-oxa-4- azaspiro[2.5]octane-
4-carboxylate 465.1 (M+), 467.1 (M + 2) .sup.a Additional Br.sub.2
(0.5 equivalents) was necessary
[0432] Intermediate L16
##STR00082##
2-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)-2-methylpropanenitril-
e
[0433] A solution of
2-(4-(5-aminopyrazin-2-yl)-1H-pyrazol-1-yl)-2-methylpropanenitrile
(Intermediate Y16; 77.0 mg, 0.337 mmol) in CHCl.sub.3 (3.4 mL) was
treated with pyridine (30.1 .mu.L, 0.371 mmol). The resulting
solution was cooled to 0.degree. C. and then treated with Br.sub.2
(18.2 .mu.L, 0.354 mmol). The reaction mixture was stirred 16 h at
ambient temperature prior to quenching with 10%
Na.sub.2S.sub.2O.sub.3(aq) (10 mL). The resulting biphasic mixture
was diluted with 4:1 DCM:iPrOH (50 mL) and washed with water
(2.times.25 mL). The combined organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The resulting crude residue was purified by silica
chromatography (1-25% DCM/MeOH as the gradient eluent) to afford
the title compound (79.3 mg, 77% yield). MS (apci) m/z=307.0
[(M+H)+2], 308.9 (M+H), with Br pattern.
[0434] The following intermediates shown in Table L16 were prepared
according the method used for the synthesis of Intermediate L16
from the appropriate starting materials prepared as described
herein. Reaction progression in each was followed by LCMS and
reaction time was adjusted as necessary. All compounds were
purified by silica chromatography according to the method for
isolating Intermediate L16 using the appropriate gradient
eluent.
TABLE-US-00010 TABLE L16 Intermediate MS (apci) # Structure Name
m/z L14 ##STR00083## 1-((4-(5-amino- 6-bromopyrazin-
2-yl)-1H-pyrazol-1- yl)methyl)cyclopropane- 1-carbonitrile 320.9
[(M + H) + 2] 319.0 (M + H) (with bromine pattern) L15 ##STR00084##
1-(4-(5-amino-6- bromopyrazin-2- yl)-1H-pyrazol- 1-yl)-4,4,4-
trifluorobutan-2-ol 367.9 [(M + H) + 2] 366 (M + H) (with bromine
pattern) L18 ##STR00085## 2-(4-(5-amino-6- bromopyrazin-2-
yl)-1H-pyrazol- 1-yl)-2-methylpropan- 1-ol 314.0 [(M + H) + 2] 312
(M + H) (with bromine pattern)
[0435] Intermediate L20
##STR00086##
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-amine
[0436] A solution of 5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-amine
(Intermediate Y20; 845.5 mg, 4.854 mmol) in CHCl.sub.3 (25 mL) was
treated with pyridine (431.8 .mu.L, 5.339 mmol). The resulting
solution was cooled to 0.degree. C. then treated with Br.sub.2
(261.1 .mu.L, 5.096 mmol). The reaction mixture was stirred 16 h at
ambient temperature prior to quenching with 10%
Na.sub.2S.sub.2O.sub.3(aq) (10 mL). The resulting biphasic mixture
was extracted with CHCl.sub.3 (2.times.100 mL). The combined
organic extracts washed with 10% Na.sub.2S.sub.2O.sub.3(aq) (25
mL), then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum. The resulting crude residue was purified
by silica chromatography (1-25% DCM/MeOH as the gradient eluent) to
afford the title compound (500.5 mg, 41% yield). MS (apci)
m/z=254.9 [(M+H)+2], 252.9 (M+H), with Br pattern.
[0437] Intermediate L21
##STR00087##
tert-butyl
4-(4-(6-amino-5-bromopyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te
[0438] The title compound was prepared (622.4 mg, 45% yield)
according to the method described for Intermediate L20, using
tert-butyl
4-(4-(6-aminopyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
(Intermediate Y21) in place of
5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-amine with excess
bromine/pyridine (1.6 eq each) and a gradient eluent system of
10-90% DCM/EtOAc in the silica chromatography. MS (apci) m/z=424
[(M+H)+2], 422.0 (M+H) with Br pattern.
[0439] Intermediate L22
##STR00088##
3-bromo-5-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-
-amine
[0440] Step 1: Tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 2.0 g, 4.7 mmol) was treated with TFA (5 mL)
and stirred at ambient temperature. After 30 minutes, the TFA was
removed in vacuo and the residue was treated with 4N HCl in dioxane
(50 mL) to form the HCl salt. The resulting mixture was
concentrated in vacuo and the residue was dried under high vacuum
to a constant weight to provide
3-bromo-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-amine
dihydrochloride (2.1 g, 5.3 mmol, 112% yield) as a white solid. MS
(apci) m/z=325 [(M+H)+2], 323.0 (M+H) with Br pattern.
[0441] Step 2: 3-Bromo-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)
pyrazin-2-amine dihydrochloride (1.9 g, 4.797 mmol) was dissolved
in DMF (100 mL) and treated with K.sub.2CO.sub.3 (2.652 g, 19.19
mmol). The mixture was cooled to 0.degree. C. and treated dropwise
with 1-bromo-2-methoxyethane (0.4508 ml, 4.797 mmol) while
maintaining the internal temperature at 0.degree. C., and then
allowed to warm to ambient temperature while stirring for 72 hours.
The reaction mixture was poured into ice water (1.0 L) and
extracted with 5% IPA in DCM. The organics were washed with brine,
then, dried over MgSO.sub.4, filtered and concentrated in vacuo.
The crude product was purified by flash chromatography (2-15% MeOH
in DCM with 2% NH.sub.4OH) to provide the title compound (1.2 g,
3.147 mmol, 65.62% yield) as an off white solid. MS (apci)
m/z=383.1 [(M+H)+2], 381 (M+H) with Br pattern.
[0442] Intermediate X2
##STR00089##
6-chloro-2-(3-methoxy-5-(trifluoromethoxy)phenyl)pyridazin-3
(2H)-one
[0443] A mixture of 6-chloropyridazin-3(2H)-one (0.125 g, 0.958
mmol), (3-methoxy-5-(trifluoromethoxy)phenyl)boronic acid (0.339 g,
1.44 mmol), Cu(OAc).sub.2 (0.0348 g, 0.192 mmol) and pyridine
(0.155 mL, 1.92 mmol) in DCM (9.58 mL) was stirred overnight at
ambient temperature. The mixture was diluted with DCM and washed
with water and brine then dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by silica chromatography to afford the title compound
(307 mg, 82% yield). MS (apci) m/z=323.0 [(M+H)+2], 321.0 (M+H)
with Cl pattern.
[0444] The following intermediates, shown in Table X2 were prepared
according the method used for the synthesis of Intermediate X2
using the appropriate arylboronic acid starting materials. Reaction
progression in each was followed by LCMS and reaction time was
adjusted as necessary. All compounds were purified by silica
chromatography according to the method used to isolate Intermediate
X2.
TABLE-US-00011 TABLE X2 Intermediate Structure Name Spectral Data
X1 .sup.a ##STR00090## 6-chloro-2-(3- methoxyphenyl)pyridazin-
3(2H)-one .sup.1H NMR (DMSO-d.sub.6) .delta. 7.60 (d, 1H), 7.38 (t,
1H), 7.12 (d, 1H), 7.07 (m, 2H), 7.00 (m, 1H), 3.75 (s, 3H) X5
##STR00091## 6-chloro-2-(3- isopropoxy-5- methoxyphenyl)pyridazin-
3(2H)-one MS (apci) m/z = 297.1 [(M + H) + 2], 295.0 (M + H) with
Cl pattern X7* ##STR00092## 6-chloro-2-(o- tolyl)pyridazin-3(2H)-
one .sup.1H NMR (CDCl.sub.3) .delta. 7.30 (m, 4H), 7.04 (d, 1H),
2.19 (s, 3H) .sup.a used 10 times the amount of Cu(OAc).sub.2 used
for preparing Intermediate X2 but otherwise the procedure was as
described for Intermediate X2
[0445] Intermediate X3
##STR00093##
6-chloro-2-(3-ethoxy-5-(trifluoromethoxy)phenyl)pyridazin-3
(2H)-one
[0446]
6-chloro-2-(3-ethoxy-5-(trifluoromethoxy)phenyl)pyridazin-3(2H)-one
was made according to the procedure of Intermediate X2 substituting
(3-methoxy-5-(trifluoromethoxy)phenyl)boronic acid for
(3-ethoxy-5-(trifluoromethoxy)phenyl)boronic acid.
[0447] Intermediate X4
##STR00094##
6-chloro-2-(3-methoxy-5-(trifluoromethyl)phenyl)pyridazin-3
(2H)-one
[0448]
6-chloro-2-(3-methoxy-5-(trifluoromethyl)phenyl)pyridazin-3(2H)-one
was made according to the procedure for Intermediate X2,
substituting (3-methoxy-5-(trifluoromethoxy)phenyl)boronic acid for
(3-methoxy-5-(trifluoromethyl)phenyl)boronic acid.
[0449] Intermediate X6
##STR00095##
6-chloro-2-(2-chloro-3-methoxyphenyl)pyridazin-3 (2H)-one
[0450] 6-chloro-2-(2-chloro-3-methoxyphenyl)pyridazin-3 (2H)-one
was made according to the procedure for Intermediate X2,
substituting (3-methoxy-5-(trifluoromethoxy)phenyl)boronic acid for
(2-chloro-3-methoxyphenyl)boronic acid.
[0451] Intermediate X8
##STR00096##
6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0452] A solution of 6-chloropyridazin-3(2H)-one (506.5 mg, 3.880
mmol) in DCM (38 mL) was treated with 3,5-dimethoxyphenylboronic
acid (776.8 mg, 4.268 mmol), Cu(OAc).sub.2 (1410 mg, 7.760 mmol),
and pyridine (627.7 .mu.L, 7.760 mmol). The resulting mixture was
stirred open to the atmosphere for 60 h at ambient temperature. The
reaction mixture was filtered, and the filtrate was concentrated
under vacuum. The resulting crude residue was purified by silica
chromatography (5-60% DCM/EtOAc as the gradient eluent) to afford
the title compound (560 mg, 54% yield). MS (apci) m/z=269.0
[(M+H)+2], 267.0 (M+H), with Cl pattern.
[0453] The following intermediates shown in Table X8 were prepared
according the method used for the synthesis of Intermediate X8
using the appropriate arylboronic acid starting materials. Reaction
progression in each was followed by LCMS and reaction time was
adjusted as necessary. All compounds were purified by silica
chromatography according to the method used to isolate Intermediate
X8.
TABLE-US-00012 TABLE X8 Intermediate Structure Name Spectral Data
X9 ##STR00097## 6-chloro-2-(3,4- dimethoxyphenyl)pyridazin-
3(2H)-one MS (apci) m/z = 269.0 [(M + H) + 2], 267.0 (M + H), with
Cl pattern X16 ##STR00098## methyl 3-(3-chloro- 6-oxopyridazin-
1(6H)-yl)benzoate .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
8.15-8.14 (m, 1H), 8.03-8.00 (m, 1H), 7.89-7.86 (m, 1H), 7.69-7.65
(m, 2H), 7.21- 7.18 (d, 1H), 3.89 (s, 3H)
[0454] Intermediate X10
##STR00099##
6-chloro-2-(5-methoxy-2-methylphenyl)pyridazin-3(2H)-one
[0455] A solution of 6-chloropyridazin-3(2H)-one (811.1 mg, 6.214
mmol) in DCM (31 mL) was treated with
(5-methoxy-2-methylphenyl)boronic acid (1031 mg, 6.214 mmol),
Cu(OAc).sub.2 (2257 mg, 12.43 mmol), and pyridine (1005 .mu.L,
12.43 mmol). The resulting mixture was stirred open to the
atmosphere for 16 h at ambient temperature. The reaction mixture
was filtered, and the filtrate was concentrated under vacuum. The
resulting crude residue was purified by C18 reverse phase
chromatography (5-95% water/ACN with 0.1% TFA as the gradient
eluent) to afford the title compound as a TFA salt. The TFA salt
was dissolved in 4:1 DCM:iPrOH (100 mL) and extracted with
saturated NaHCO.sub.3(aq) (1.times.25 mL). The organic extracts
were dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum cleanly affording the title compound (251
mg, 16% yield). MS (apci) m/z=253.0 [(M+H)+2], 251.0 (M+H) with Cl
pattern.
[0456] Intermediate X11
##STR00100##
6-chloro-2-(1,5-dimethyl-1H-indazol-4-yl)pyridazin-3(2H)-one
[0457] The title compound was prepared (52 mg, 12% yield) according
to the method described for Intermediate X10, using
1,5-dimethyl-1H-indazole-4-boronic acid in place of
(5-methoxy-2-methylphenyl)boronic acid. MS (apci) m/z=277.0
[(M+H)+2], 275.0 (M+H) with Cl pattern.
[0458] Intermediate X12
##STR00101##
3-(3-chloro-6-oxopyridazin-1(6H)-yl)-5-methoxybenzonitrile
Step 1: Preparation of (3-cyano-5-methoxyphenyl)boronic acid
[0459] A mixture of
3-methoxy-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzonitrile
(0.450 g, 1.74 mmol), sodium periodate (1.11 g, 5.21 mmol) and 1 M
CH.sub.3COONH.sub.4(aq) (3.47 mL, 3.47 mmol) in acetone (7 mL) was
stirred 3 h at ambient temperature. The reaction was quenched with
4 M HCl.sub.(aq) (1 mL) then stirred for 20 min. The mixture was
then diluted with EtOAc and extracted with water and brine, then
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum to afford the title compound (0.300 g, 98% yield).
Step 2: Preparation of
3-(3-chloro-6-oxopyridazin-1(6H)-yl)-5-methoxybenzonitrile
[0460] A mixture of 6-chloropyridazin-3(2H)-one (0.200 g, 1.53
mmol), (3-cyano-5-methoxyphenyl)boronic acid (0.298 g, 1.69 mmol),
Cu(OAc).sub.2 (0.0557 g, 0.306 mmol) and pyridine (0.273 mL, 3.37
mmol) in DCM (9.58 mL) was stirred overnight at ambient
temperature. The mixture was then diluted with DCM and extracted
with water and brine, then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting crude residue was purified by silica chromatography to
afford the title compound (197 mg, 49% yield). .sup.1H NMR
(CDCl.sub.3) .delta. 7.60 (t, 1H), 7.49 (t, 1H), 7.28 (d, 1H), 7.18
(m, 1H), 7.40 (d, 1H), 3.88 (s, 3H).
[0461] Intermediate X13
##STR00102##
6-chloro-2-(2-fluoro-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of (2-fluoro-3,5-dimethoxyphenyl)hydrazine
[0462] A cold (0.degree. C.) solution of
2-fluoro-3,5-dimethoxyaniline (1.00 g, 5.84 mmol) in 12.5 M
HCl.sub.(aq) (7.01 mL, 87.6 mmol) was slowly treated with
NaNO.sub.2(s) (0.605 g, 8.76 mmol) then stirred for 1 h at ambient
temperature. The resulting reaction mixture then was treated with
SnCl.sub.2.H.sub.2O (2.64 g, 11.7 mmol), and stirred overnight at
ambient temperature. The reaction mixture then was filtered,
washing the solids with water. The filtrate was cooled to 0.degree.
C. and slowly basified with the addition of NaOH pellets. The
resulting mixture was extracted with ethyl acetate (5.times.250
mL), and the combined organic extracts were washed with brine, then
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum to afford the title compound (724 mg, 67% yield). MS
(apci) m/z=187.1 (M+H). This material was used without purification
in the subsequent step.
Step 2: Preparation of
1-(2-fluoro-3,5-dimethoxyphenyl)-1,2-dihydropyridazine-3,6-dione
[0463] A solution of furan-2,5-dione (0.381 g, 3.89 mmol) and
(2-fluoro-3,5-dimethoxyphenyl)hydrazine (from step 1; 0.724 g, 3.89
mmol) in EtOH (absolute; 19.4 mL) was heated for 1 day at
95.degree. C. then treated with 6 N HCl in iPrOH (2.5 mL). After 3
h the reaction mixture was concentrated under vacuum. The resulting
residue was suspended in DCM and the insoluble material was removed
by filtration. The filtrate then was concentrated under vacuum, and
the residue was purified by silica chromatography (0-100%
EtOAc/hexanes as the gradient eluent) to cleanly afford the title
compound (140.5 mg, 14% yield). MS (apci) m/z=267.0 (M+H).
Step 3: Preparation of
6-chloro-2-(2-fluoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0464] A solution of
1-(2-fluoro-3,5-dimethoxyphenyl)-1,2-dihydropyridazine-3,6-dione
(140 mg, 0.526 mmol) in POCl.sub.3 (490 .mu.L, 5.26 mmol) was
heated at 85.degree. C. for 1 h. The reaction mixture was then
concentrated under vacuum, and the resulting residue was
partitioned between EtOAc and saturated NaHCO.sub.3(aq). The phases
were separated and treated independently. The aqueous extracts were
washed with EtOAc (2x) and the organic extracts from the wash were
combined with the original organic extract. The combined organic
extracts were washed with brine then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and the concentrated under vacuum.
The resulting residue was purified by silica chromatography (0-100%
EtOAc/hexanes as the gradient eluent) to afford the title compound
(94 mg, 63% yield). MS (apci) m/z=285.0 (M+H).
[0465] Intermediate X14
##STR00103##
6-chloro-2-(2-fluoro-5-methoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of
1-(2-fluoro-5-methoxyphenyl)-1,2-dihydropyridazine-3,6-dione
[0466] A solution of furan-2,5-dione (255 mg, 2.60 mmol) and
(2-fluoro-3,5-dimethoxyphenyl)hydrazine hydrochloride (500 mg, 2.60
mmol) in EtOH (absolute; 152 mL, 2.60 mmol) was heated for 1 day at
95.degree. C. The reaction mixture was concentrated under vacuum,
and the resulting residue was purified by silica chromatography
(0-100% EtOAc/hexanes as the gradient eluent) to cleanly afford the
title compound (300 mg, 49% yield). MS (apci) m/z=237.0 (M+H).
Step 2: Preparation of
6-chloro-2-(2-fluoro-5-methoxyphenyl)pyridazin-3(2H)-one
[0467] A solution of
1-(2-fluoro-5-methoxyphenyl)-1,2-dihydropyridazine-3,6-dione (90
mg, 0.38 mmol) in POCl.sub.3 (355 .mu.L, 3.8 mmol) was heated at
85.degree. C. for 1 h. The reaction mixture was then concentrated
under vacuum, and the resulting residue was partitioned between
EtOAc and saturated NaHCO.sub.3(aq). The phases were separated and
treated independently. The aqueous extracts were washed with EtOAc
(2x), and the organic extracts from the wash were combined with the
original organic extract. The combined organic extracts were washed
with brine then dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and the concentrated under vacuum. The resulting residue was
purified by silica chromatography (0-100% EtOAc/hexanes as the
gradient eluent) to afford the title compound (38 mg, 39% yield).
MS (apci) m/z=255.0 (M+H).
[0468] Intermediate X15
##STR00104##
6-chloro-2-(2-chloro-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0469] A cold (0.degree. C.) solution of
6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one (Intermediate
X8; 0.357 g, 1.34 mmol) in ACN (13.4 mL) was treated with
SO.sub.2Cl.sub.2 (0.109 mL, 1.34 mmol) and stirred for 20 min. The
resulting mixture was quenched with the addition of saturated
NaHCO.sub.3(aq). The resulting biphasic mixture was extracted with
EtOAc. The organic extracts were washed successively with water and
brine then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
the concentrated under vacuum. The resulting crude residue was
purified by silica chromatography to afford the title compound (310
mg, 77% yield). .sup.1H NMR (CDCl.sub.3) .delta. 7.29 (d, 1H), 7.04
(d, 1H), 6.58 (m, 2H), 3.90 (s, 3H), 3.81 (s, 3H).
[0470] Intermediate O2
##STR00105##
6-chloro-2-(2-chloro-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0471] A cold (0.degree. C.) solution of
6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one (2.496 g, 9.360
mmol) in ACN (93.60 mL, 9.360 mmol) was treated with sulfuryl
dichloride (1.484 mL, 18.25 mmol) and stirred for 1 hr. The
resulting mixture was quenched with the addition of saturated
NaHCO.sub.3(aq). The resulting biphasic mixture was extracted with
DCM. The organic extracts were washed successively with water and
brine then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
then concentrated in vacuo to afford the title compound (3.1 g, 99%
yield). This material was of sufficient purity to be used directly
without further purification. .sup.1H NMR (CDCl.sub.3) .delta. 7.29
(d, 1H), 7.04 (d, 1H), 6.65 (s, 1H), 3.94 (s, 6H).
[0472] Intermediate X17
##STR00106##
6-chloro-4-methylpyridazin-3(2H)-one
Step 1: Preparation of 6-chloro-3-methoxy-4-methylpyridazine
[0473] A solution of 2,2,6,6-tetramethylpiperidine (12.9 ml, 76.1
mmol) in THF (100 mL) was sparged with N.sub.2(g) then cooled to
-78.degree. C. The -78.degree. C. solution was treated slowly with
2.5 M n-butyllithium in hexane (30.4 mL, 76.1 mmol) then warmed to
0.degree. C. and stirred for 1 h. The resulting reaction mixture
was cooled to -78.degree. C. then treated with a 0.46 M solution of
3-Chloro-6-methoxypyridazine in THF (75 mL, 34.6 mmol). After
stirring at -78.degree. C. for 1 h, the reaction mixture was
treated with iodomethane (4.74 mL, 76.1 mmol), and stirred for an
additional 30 min at -78.degree. C. The reaction mixture was
quenched with saturated NH.sub.4Cl.sub.(aq) (50 mL), warmed to
ambient temperature, diluted with water (50 mL) and extracted with
EtOAc (2.times.250 mL). The combined organic extracts were washed
with brine (1.times.50 mL), dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
afford the title compound (3.31 g, 60% yield). MS (apci) m/z=159.0
(M+H).
Step 2: Preparation of 6-chloro-4-methylpyridazin-3(2H)-one
[0474] A solution of 6-chloro-3-methoxy-4-methylpyridazine (3.31 g,
20.9 mmol) in 4:1 dioxane:water (100 mL) was treated with 12.0 M
HCl.sub.(aq) (1.91 mL, 23.0 mmol) and stirred for 60 h at
60.degree. C. The reaction mixture was concentrated under vacuum,
and the resulting crude residue was purified by silica
chromatography (1-30% DCM/MeOH with 2% NH.sub.4OH as the gradient
eluent) to afford the title compound (2.99 g, 99% yield). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 13.03 (s, 1H), 7.44 (s, 1H),
2.05 (s, 3H).
[0475] Intermediate X18
##STR00107##
6-chloro-2-(3,5-dimethoxyphenyl)-4-methylpyridazin-3 (2H)-one
[0476] A solution of 6-chloro-4-methylpyridazin-3-ol (Intermediate
X17; 500 mg, 3.46 mmol) in DCM (20.3 mL) and pyridine (1 mL, 3.46
mmol) was treated with (3,5-dimethoxyphenyl)boronic acid (1.26 g,
6.92 mmol), Cu(OAc).sub.2 (1.26 g, 6.92 mmol), and pyridine 1-oxide
(1.32 g, 13.8 mmol). The resulting mixture was stirred open to the
atmosphere overnight at ambient temperature. The reaction mixture
was diluted with DCM (100 mL) and filtered. The filtrate was washed
with water (2.times.30 mL), and the organics were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The crude residue was precipitated from MeOH to cleanly
afford the title compound (780 mg, 80%). MS (apci) m/z=281.1 (M+H),
283.0 [(M+H)+2] (with Cl pattern).
[0477] Intermediate X19
##STR00108##
methyl
3-(3-chloro-5-methyl-6-oxopyridazin-1(6H)-yl)-4-methylbenzoate
[0478] A solution of 6-chloro-4-methylpyridazin-3-ol (Intermediate
X17; 0.50 g, 3.4 mmol) in DCM (20 mL) was treated with
(5-(methoxycarbonyl)-2-methylphenyl)boronic acid (1.0 g, 5.2 mmol),
Cu(OAc).sub.2 (1.2 g, 6.9 mmol), pyridine 1-oxide (327 mg, 3.44
mmol) and pyridine (1.1 g, 14 mmol). The resulting mixture was
stirred at ambient temperature open to the atmosphere for one
overnight. The reaction mixture was diluted with DCM (100 mL) and
washed with water (2.times.30 mL). The organic extracts were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The crude residue was purified by silica gel flash
chromatography (2-55% EtOAc/hexane as the gradient eluent) to
afford the title compound (2.99 g, 99% yield). MS (apci) m/z=293.0
(M+H), 295.0 [(M+H)+2] (with Cl pattern).
[0479] Intermediate X20
##STR00109##
4-bromo-6-chloropyridazin-3 (2H)-one
[0480] A solution of 6-chloropyridazin-3-ol (5.01 g, 38.38 mmol),
KBr (13.70 g, 115.1 mmol), and KOAc (5.650 g, 57.57 mmol) in water
(80 mL) was stirred for 15 min then treated with Br.sub.2 (5.90 mL,
115 mmol). The resulting mixture was stirred under an atmosphere of
N.sub.2(g) for 2 h at 90.degree. C. After cooling to ambient
temperature the reaction mixture was quenched with 10%
Na.sub.2S.sub.2O.sub.3(aq) (100 mL). The resulting biphasic
suspension was filtered, and the filter cake was successively
rinsed with water (100 mL) then 10% Na.sub.2S.sub.2O.sub.3(aq) (100
mL). The solid filter cake was dried under high vacuum for 16 h to
cleanly afford the title compound (6.14 g, 76% yield). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 13.52 (s, 1H), 8.20 (s, 1H).
[0481] Intermediate X21
##STR00110##
4-bromo-6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0482] A mixture of 4-bromo-6-chloropyridazin-3(2H)-one
(Intermediate X20; 0.504 g, 2.41 mmol),
(3,5-dimethoxyphenyl)boronic acid (0.482 g, 2.65 mmol),
Cu(OAc).sub.2 (0.0874 g, 0.481 mmol) and pyridine (0.389 mL, 4.81
mmol) in DCM (24.1 mL) was stirred overnight at ambient temperature
The mixture was then diluted with DCM and extracted with water. The
organic extracts were washed with brine, then dried aver anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting crude residue was purified by silica chromatography to
afford the title compound (0.574 g, 69% yield). .sup.1H NMR
(CDCl.sub.3) .delta. 7.69 (s, 1H), 6.73 (d, 2H), 6.51 (t, 1H), 3.81
(s, 6H).
[0483] Intermediate X22
##STR00111##
6-chloro-4-cyclobutyl-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0484] A cold (0.degree. C.) solution of
4-bromo-6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(Intermediate X21; 413.7 mg, 1.197 mmol) in THF (12 mL) was treated
with 0.5 M cyclobutylmagnesium chloride hexane (3591 .mu.L, 1.796
mmol) and stirred for 1 h at 0.degree. C. The reaction was quenched
with the addition of water (25 mL) and the volatiles were removed
under vacuum. The remaining aqueous mixture was diluted with EtOAc
(100 mL) and washed successively with water (2.times.25 mL) and
brine (25 mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting residue was purified by silica chromatography (5-60%
Hexanes/EtOAc as the gradient eluent) to afford the title compound
(283 mg, 72% yield). MS (apci) m/z=311.0 [(M+H)+2], 309.0 (M+H)
with Cl pattern.
[0485] The following intermediates shown in Table X22 were prepared
according to the method used for the synthesis of Intermediate X22
using the appropriate alkylmagnesium halide starting materials and
4-bromo-6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one.
Reaction progression in each was followed by LCMS and reaction time
was adjusted as necessary. All compounds were purified using a
method similar to that used to isolate Intermediate X22 utilizing
the appropriate gradient eluent.
TABLE-US-00013 TABLE X22 Intermediate Structure Name MS (apci) m/z
X23 ##STR00112## 6-chloro-2-(3,5- dimethoxyphenyl)-4-
isopropylpyridazin-3(2H)- one 311.0 [(M + H) + 2], 309.0 (M + H)
with Cl pattern X24 ##STR00113## 6-chloro-4-cyclopropyl-2- (3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 309.0 [(M + H) + 2], 307.0 (M
+ H) with Cl pattern
[0486] Intermediate X25
##STR00114##
6-chloro-2-(3,5-dimethoxyphenyl)-4-isobutylpyridazin-3 (2H)-one
[0487] A cold (0.degree. C.) solution of
4-bromo-6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(Intermediate X21; 0.150 g, 0.434 mmol) in THF (2.89 mL) was
treated with 2 M isobutylmagnesium bromide in Et.sub.2O (0.434 mL,
0.868 mmol). The reaction was stirred overnight at ambient
temperature and then worked up with EtOAc and water. The organics
were washed with brine, then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting residue was purified by silica chromatography to afford
the title compound (0.035 g, 25% yield).
[0488] The following intermediates, shown in Table X25 were
prepared according the method used for the synthesis of
Intermediate X25 using the appropriate alkylmagnesium halide
starting materials and
4-bromo-6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one. All
compounds were purified by silica chromatography utilizing the
appropriate gradient.
TABLE-US-00014 TABLE X25 Intermediate Structure Name X26
##STR00115## 6-chloro-2-(3,5-dimethoxyphenyl)-4-
ethylpyridazin-3(2H)-one X27 ##STR00116##
6-chloro-2-(3,5-dimethoxyphenyl)-4- propylpyridazin-3(2H)-one
[0489] Intermediate X28
##STR00117##
6-chloro-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3
(2H)-one
[0490] Sulfuryl chloride (0.152 mL, 1.88 mmol) was added to
6-chloro-2-(3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one
(Intermediate X18; 0.264 g, 0.940 mmol) in ACN (6.27 mL) at
0.degree. C. This was then stirred at room temp for 20 min. The
mixture was then quenched with saturated aqueous Na.sub.2CO.sub.3.
The mixture was partitioned between EtOAc and water. The combined
organic layers were washed with water and brine, dried over
Na.sub.2SO.sub.4, and concentrated. The residue was purified on a
silica column using Hexanes:EtOAc (10-90%) to give
6-chloro-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one
(0.198 g, 0.566 mmol, 60.2% yield) MS (apci) m/z=349.0 (M+H).
[0491] Intermediate X29
##STR00118##
6-chloro-2-(3-methylquinolin-7-yl)pyridazin-3 (2H)-one
[0492] Step 1: Preparation of (3-methylquinolin-7-yl)boronic acid:
A solution of 7-bromo-3-methylquinoline (258 mg, 1.16 mmol),
5,5,5',5'-tetramethyl-2,2'-bi(1,3,2-dioxaborinane) (656 mg, 2.90
mmol), PdCl2(dppf).dcm (47.4 mg, 0.0581 mmol), and KOAc (342 mg,
3.49 mmol) in dioxane (5809 .mu.L, 1.16 mmol) was sparged with
N.sub.2(g) for 5 min at ambient temperature and then heated at
90.degree. C. overnight. After cooling to ambient temperature, the
reaction mixture was partitioned between EtOAc and water. The
combined organic extracts were washed with brine, dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to afford the title compound with higher than expected mass,
but assumed with quantitative yield (217 mg, 100% yield). MS (apci)
m/z=188.1 (M+H).
[0493] Step 2: Preparation of:
6-chloro-2-(3-methylquinolin-7-yl)pyridazin-3(2H)-one: A mixture of
6-chloropyridazin-3(2H)-one (0.140 g, 1.07 mmol),
(3-methylquinolin-7-yl)boronic acid, (0.221 g, 1.18 mmol), Cu(OAc)2
(0.0390 g, 0.215 mmol) and pyridine (0.191 ml, 2.36 mmol) in DCM
(10.7 mL, 1.07 mmol) was stirred at room temperature overnight. The
reaction mixture was partitioned between EtOAc and water. The
organic extracts were washed with brine, then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting residue was purified by silica chromatography to afford
the title compound (25 mg, 8.6% yield). MS (apci) m/z=274.0
[(M+H)+2], 272.0 (M+H) with Cl pattern.
[0494] Intermediate X30
##STR00119##
Methyl 3-(3-chloro-5-methyl-6-oxopyridazin-1
(6H)-yl)-4-methylbenzoate
[0495] A solution of 6-chloro-4-methylpyridazin-3-ol (0.5 g, 3.5
mmol) in dichloromethane (20 mL, 3.4 mmol) was treated with
(5-(methoxycarbonyl)-2-methylphenyl)boronic acid (1 g, 5.15 mmol),
copper(II) acetate (1.25 g, 6.87 mmol), pyridine 1-oxide (33 mg,
3.44 mmol), and pyridine (1.1 g, 13.75 mmol). The resulting mixture
was stirred overnight at RT. The mixture was diluted with DCM (100
mL) and washed with water (2.times.30 mL). The organic layer was
separated, dried (Na.sub.2SO.sub.4), filtered and concentrated in
vacuo. The residue was purified by flash chromatography on silica
gel (Redi Sep 80 g) eluting with 2-55% EtOAc/hexane to provide
methyl
3-(3-chloro-5-methyl-6-oxopyridazin-1(6H)-yl)-4-methylbenzoate (430
mg, 43% yield) as a solid. LCMS (APCI+) m/z 293.0 (M+1); retention
time=4.086 min.
[0496] Intermediate R1
##STR00120##
2-(3-methoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrida-
zin-3(2H)-one
[0497] A solution of
6-chloro-2-(3-methoxyphenyl)pyridazin-3(2H)-one (Intermediate X1;
104 mg, 0.439 mmol) in dioxane (6 mL) was treated with
bis(pinacolato)diboron (123 mg, 0.483 mmol), Pd(OAc).sub.2 (10.8
mg, 0.0483 mmol), X-Phos (34.6 mg, 0.0725 mmol), and KOAc (129 mg,
1.32 mmol). The mixture was sparged with Ar.sub.(g), then sealed
and stirred for 16 h at 100.degree. C. After cooling to ambient
temperature, the reaction mixture was filtered, and the filter cake
was washed with EtOAc (50 mL). The filtrate was concentrated under
vacuum to afford the title compound (157 mg, 109% crude yield).
This material was used directly without further purification.
[0498] Intermediate R2
##STR00121##
2-(3-methoxy-5-(trifluoromethoxy)phenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dio-
xaborolan-2-yl)pyridazin-3 (2H)-one
[0499] A mixture of
6-chloro-2-(3-methoxy-5-(trifluoromethoxy)phenyl)pyridazin-3(2H)-one
(Intermediate X2; 0.253 g, 0.789 mmol), bis(pinacolato)diboron
(0.220 g, 0.868 mmol), Pd(OAc).sub.2 (0.0177 g, 0.0789 mmol),
X-Phos (0.0564 g, 0.118 mmol), and KOAc (0.232 g, 2.37 mmol) in
dioxane (2.63 mL) was sparged with Ar.sub.(g) for 5 min at ambient
temperature then stirred for 1 h at 100.degree. C. After cooling to
ambient temperature, the reaction mixture was partitioned between
EtOAc and water. The organic extracts were washed with brine, then
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum to afford the title compound (320 mg, 98% yield). MS
(apci) m/z=287.0 (M-B(OR).sub.2+H). This material was of sufficient
purity to be used directly without further purification.
[0500] The following intermediates shown in Table R2 were prepared
according the method used for the synthesis of Intermediate R2
using the appropriate 6-chloro-2-(Aryl)pyridazin-3(2H)-one starting
materials (Intermediates X3-X7, X11-X12, X15). Reaction progression
for each was followed by LCMS and reaction time was adjusted as
necessary.
TABLE-US-00015 TABLE R2 MS (apci) Intermediate Structure Name m/z
R3 ##STR00122## 2-(3-ethoxy-5- (trifluoromethoxy)phenyl)-
6-(4,4,5,5- tetramethyl-1,3,2- dioxaborolan-2-
yl)pyridazin-3(2H)-one 301.1 (M--B(OR).sub.2 + H) R4 ##STR00123##
2-(3-methoxy-5- (trifluoromethyl)phenyl)- 6-(4,4,5,5-
tetramethyl-1,3,2- dioxaborolan-2- yl)pyridazin-3(2H)-one 271.1
(M--B(OR).sub.2 + H) R5 ##STR00124## 2-(3-isopropoxy-5-
methoxyphenyl)-6- (4,4,5,5-tetramethyl- 1,3,2-dioxaborolan-2-
yl)pyridazin-3(2H)-one 261.1 (M--B(OR).sub.2 + H) R6 ##STR00125##
2-(2-chloro-3- methoxyphenyl)-6- (4,4,5,5-tetramethyl-
1,3,2-dioxaborolan-2- yl)pyridazin-3(2H)-one 237 (M--B(OR).sub.2 +
H) R12 ##STR00126## 3-methoxy-5-(6-oxo-3- (4,4,5,5-tetramethyl-
1,3,2-dioxaborolan-2- yl)pyridazin-1(6H)- yl)benzonitrile 226.1
(M--B(OR).sub.2 + H) R15 ##STR00127## 2-(2-chloro-3,5-
dimethoxyphenyl)-6- (4,4,5,5-tetramethyl- 1,3,2-dioxaborolan-2-
yl)pyridazin-3(2H)-one 267 (M--B(OR).sub.2 + H) R16 ##STR00128##
2-(2,6-dichloro-3,5- dimethoxyphenyl)-4- methyl-6-(4,4,5,5-
tetramethyl-1,3,2- dioxaborolan-2- yl)pyridazin-3(2H)-one 315
(M--B(OR).sub.2 + H) R17 ##STR00129## 2-(2,6-dichloro-3,5-
dimethoxyphenyl)-6- (4,4,5,5-tetramethyl- 1,3,2-dioxaborolan-2-
yl)pyridazin-3(2H)-one 301 (M--B(OR).sub.2 + H)
[0501] Intermediate R7
##STR00130##
6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-2-(o-tolyl)pyridazin-3(2H-
)-one
[0502]
6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-2-(o-tolyl)pyridazi-
n-3 (2H)-one was made according the procedure of Intermediate R2,
substituting Intermediate X2 with Intermediate X7.
[0503] Intermediate R8
##STR00131##
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3 (2H)-one
[0504] A solution of 6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3
(2H)-one (Intermediate X8; 47.9 mg, 0.180 mmol) in dioxane (1.8 mL)
was treated with bis(pinacolato)diboron (50.2 mg, 0.198 mmol),
Pd(OAc).sub.2 (4.03 mg, 0.0180 mmol), X-Phos (12.8 mg, 0.0269
mmol), and KOAc (52.9 mg, 0.539 mmol). The mixture was sparged with
Ar.sub.(g), then sealed and stirred for 16 h at 100.degree. C.
After cooling to ambient temperature, the reaction mixture was
diluted with 4:1 DCM:iPrOH (25 mL) then extracted with water
(2.times.10 mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
afford the title compound (64 mg, 99% yield). MS (apci) m/z=233.1
(desB(OR).sub.2 M+H). This material was of sufficient purity to be
used directly without further purification.
[0505] The following intermediates shown in Table R8, were prepared
according the method used for the synthesis of Intermediate R8
using the appropriate 6-chloro-2-(Aryl)pyridazin-3(2H)-one starting
materials (Intermediates X9, X10). Reaction progression in each was
followed by LCMS and reaction time was adjusted as necessary.
TABLE-US-00016 TABLE R8 Inter- mediate Structure Name MS (apci) m/z
= R9 ##STR00132## 2-(3,4-dimethoxyphenyl)- 6-(4,4,5,5-tetramethyl-
1,3,2-dioxaborolan-2- yl)pyridazin-3(2H)-one 277.0 (B(OH).sub.2
M+), 233.0 (M--B(OR).sub.2 + H) R10 ##STR00133## 2-(5-methoxy-2-
methylphenyl)-6-(4,4,5,5- tetramethyl-1,3,2- dioxaborolan-2-
yl)pyridazin-3(2H)-one 261.0 (B(OH).sub.2 M+), 217.1
(M--B(OR).sub.2 + H)
[0506] Intermediate R13
##STR00134##
2-(2-fluoro-3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3 (2H)-one
[0507] A solution of
6-chloro-2-(2-fluoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(Intermediate X13; 94 mg, 0.33 mmol) in dioxane (1.2 mL) was
treated with bis(pinacolato)diboron (0.17 g, 0.66 mmol),
Pd(OAc).sub.2 (7.4 mg, 0.033 mmol), X-Phos (23.6 mg, 0.050 mmol),
and KOAc (97 mg, 0.99 mmol). The mixture was sparged with
Ar.sub.(g), then sealed and stirred for 3 h at 90.degree. C. After
cooling to ambient temperature, the reaction mixture was
partitioned between EtOAc and water. The organic extracts were
reserved and the aqueous extracts were independently washed with
EtOAc (3.times.). The combined organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to afford the title compound (124.2 mg, 75% yield). MS
(apci) m/z=251.1 (desB(OR).sub.2 M+H). This material was of
sufficient purity to be used directly without further
purification.
[0508] Intermediate R14
##STR00135##
2-(2-fluoro-5-methoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2--
yl)pyridazin-3 (2H)-one
[0509] The title compound was prepared (116.9 mg, 50% yield)
according to the method described for Intermediate R14, using
6-chloro-2-(2-fluoro-5-methoxyphenyl)pyridazin-3(2H)-one
(Intermediate X14) in place of
6-chloro-2-(2-fluoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(Intermediate X13). MS (apci) m/z=221.1 (desB(OR).sub.2 M+H). This
material was of sufficient purity to be used directly without
further purification.
[0510] Intermediate R18
##STR00136##
2-(3,5-dimethoxyphenyl)-4-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3 (2H)-one
[0511] A solution of
6-chloro-2-(3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one
(Intermediate X18; 199.4 mg, 0.7103 mmol) in dioxane (7.1 mL) was
treated with bis(pinacolato)diboron (198.4 mg, 0.7814 mmol),
Pd(OAc).sub.2 (15.95 mg, 0.07103 mmol), X-Phos (50.80 mg, 0.1066
mmol), and KOAc (209.1 mg, 2.131 mmol). The mixture was sparged
with Ar.sub.(g), then sealed and stirred for 1 h at 100.degree. C.
After cooling to ambient temperature, the reaction mixture was
diluted with 4:1 DCM:iPrOH (50 mL) then extracted with water
(2.times.25 mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
afford the title compound (264 mg, 100% yield). MS (apci) m/z=291.1
(B(OH).sub.2 M+H), 247.1 (desB(OR).sub.2 M+H). This material was of
sufficient purity to be used directly without further
purification.
[0512] Intermediate R19
##STR00137##
methyl
4-methyl-3-(5-methyl-6-oxo-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborol-
an-2-yl)pyridazin-1 (6H)-yl)benzoate
[0513] A mixture of methyl
3-(3-chloro-5-methyl-6-oxopyridazin-1(6H)-yl)-4-methylbenzoate
(Intermediate X19; 425 mg, 1.45 mmol), bis(pinacolato)diboron (553
mg, 2.18 mmol), Pd(OAc).sub.2 (32.6 mg, 0.145 mmol), X-Phos (104
mg, 0.218 mmol), and KOAc (427 mg, 4.36 mmol) in dioxane (14.5 mL)
was sparged with N.sub.2(g), then sealed and stirred for 6 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (100 mL) then extracted with
water (2.times.30 mL). The organic extracts were dried over
anhydrous MgSO.sub.4(s), filtered and concentrated under vacuum to
afford the crude title compound. MS (apci) m/z=384.1 (M+), 259.0
[(M-B(OR).sub.2)+H]. This material was of sufficient purity to be
used directly without further purification.
[0514] Intermediate R22
##STR00138##
4-cyclobutyl-2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxabo-
rolan-2-yl)pyridazin-3(2H)-one
[0515] A solution of
6-chloro-4-cyclobutyl-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(Intermediate X22; 149.6 mg, 0.4664 mmol) in dioxane (5.0 mL) was
treated with bis(pinacolato)diboron (130.3 mg, 0.5130 mmol),
Pd(OAc).sub.2 (10.47 mg, 0.04664 mmol), X-Phos (33.35 mg, 0.06996
mmol), and KOAc (137.3 mg, 1.399 mmol). The mixture was sparged
with Ar.sub.(g), then sealed and stirred for 1 h at 100.degree. C.
After cooling to ambient temperature, the reaction mixture was
diluted with 4:1 DCM:iPrOH (50 mL) and then extracted with water
(2.times.25 mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
afford the title compound (192.2 mg, 100% yield). MS (apci)
m/z=261.0 (B(OH).sub.2 M+H), 217.1 (M-B(OR).sub.2+H). This material
was of sufficient purity to be used directly without further
purification.
[0516] The following intermediates shown in Table R22 were prepared
according the method used for the synthesis of Intermediate R22
using the appropriate starting materials (Intermediates X23-X27).
Reaction progression in each was followed by LCMS and reaction time
was adjusted as necessary.
TABLE-US-00017 TABLE R22 Intermediate Structure Name R23
##STR00139## 2-(3,5-dimethoxyphenyl)-4-isopropyl-6-
(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2- yl)pyridazin-3(2H)-one
R24 ##STR00140## 4-cyclopropyl-2-(3,5-dimethoxyphenyl)-6-
(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2- yl)pyridazin-3(2H)-one
R25 ##STR00141## 2-(3,5-dimethoxyphenyl)-4-isobutyl-6-
(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2- yl)pyridazin-3(2H)-one
R26 ##STR00142## 2-(3,5-dimethoxyphenyl)-4-ethyl-6-(4,4,5,5-
tetramethyl-1,3,2-dioxaborolan-2- yl)pyridazin-3(2H)-one R27
##STR00143## 2-(3,5-dimethoxyphenyl)-4-propyl-6-
(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-
yl)pyridazin-3(2H)-one
[0517] Intermediate R28
##STR00144##
2-(2-chloro-6-fluoro-3-methoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxab-
orolan-2-yl)pyridazin-3 (2H)-one
[0518] A solution of
6-chloro-2-(2-chloro-6-fluoro-3-methoxyphenyl)pyridazin-3(2H)-one
(ArkPharm, 180 mg, 0.622 mmol), bis(pinacolato)diboron (316 mg,
1.24 mmol), palladium(II) acetate (14.0 mg, 0.062 mmol), X-PHOS (45
mg, 0.093 mmol), and potassium acetate (183 mg, 1.87 mmol). was
degassed with nitrogen, sealed, and heated to 100.degree. C.
overnight (16 hrs). The mixture was cooled to ambient temperature
then diluted with 4:1 DCM:IPA and washed with water. The organic
layer was dried over Na.sub.2SO.sub.4 and concentrated. The residue
was carried forward to next step without further purification. LCMS
(APCI+) m/z=255.0; retention time 1.60 min.
[0519] Intermediate R29
##STR00145##
2-(2,6-difluoro-3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxabo-
rolan-2-yl)pyridazin-3(2H)-one
[0520] A solution of
6-chloro-2-(2,6-difluoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(ArkPharm 163 mg, 0.539 mmol), bis(pinacolato)diboron (274 mg, 1.08
mmol), palladium(II) acetate (12.1 mg, 0.0539 mmol), X-PHOS (38.5
mg, 0.081 mmol), and potassium acetate (159 mg, 1.62 mmol) was
sparged with nitrogen, sealed, and heated to 100.degree. C. for 16
h. The reaction mixture was cooled to ambient temperature, then
diluted with 4:1 DCM:IPA, washed with water, dried over
Na.sub.2SO.sub.4, filtered through fluted filter paper and
concentrated. The residue was used without further purification in
excess in a subsequent reaction. LCMS (APCI+) m/z 269.1 (fragment:
M-pinacolboronate); Retention time=1.70 min.
[0521] Intermediate R30
##STR00146##
2-(3-methylquinolin-7-yl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-
pyridazin-3 (2H)-one
[0522] A mixture of 6-chloro-2-(3-methylquinolin-7-yl)pyridazin-3
(2H)-one (Intermediate X29; 25 mg, 0.09 mmol),
Bis(Pinacolato)diboron (70.1 mg, 0.28 mmol), Pd(OAc).sub.2 (2.1 mg,
0.01 mmol), and XPHOS (6.6 mg, 0.01 mmol) in dioxane (0.6 mL) was
sparged with Ar.sub.(g) for 5 min at ambient temperature, then
stirred for 3 h at 90.degree. C. After cooling to ambient
temperature, the reaction mixture was partitioned between EtOAc and
water. The organic extracts were washed with brine, then dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to afford the title compound (21.8 mg, 99% yield). MS (apci)
m/z=238.1 (M-B(OR).sub.2+H). This material was of sufficient purity
to be used directly without further purification.
[0523] Intermediate R31
##STR00147##
Methyl
4-methyl-3-(5-methyl-6-oxo-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborol-
an-2-yl)pyridazin-1(6H)-yl)benzoate
[0524] A glass pressure tube was charged with Intermediate X30
[methyl
3-(3-chloro-5-methyl-6-oxopyridazin-1(6H)-yl)-4-methylbenzoate]
(425 mg, 1.45 mmol), bis(pinacolato)diboron (553 mg, 2.2 mmol),
palladium (II) acetate (33 mg, 0.145 mmol), XPHOS (104 mg, 0.22
mmol), potassium acetate (427.5 mg, 4.4 mmol) and 1,4-dioxane
(14519 .mu.L, 1.45 mmol). The mixture was sparged with N2. The tube
was sealed with a Teflon screw cap and heated at 100.degree. C.
with stirring for 6 hrs. The mixture was then cooled to 0.degree.
C., diluted with 4:1 DCM:IPA (100 mL) and washed with water. The
organic layer was separated, dried over MgSO.sub.4 and concentrated
under reduced pressure. The crude product was used immediately in a
subsequent reaction.
[0525] Intermediate M1
##STR00148##
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)-N-methylbenza-
mide
Step 1: Preparation of Methyl
3-(3-(2-aminopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)benzoate
[0526] A solution of methyl
3-(3-chloro-6-oxopyridazin-1(6H)-yl)benzoate (Intermediate X16;
531.9 mg, 2.010 mmol) in 4:1 dioxane:water (15 mL) was treated with
t-butyl
3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-ylcarbamate
(643.5 mg, 2.010 mmol), Pd(PPh.sub.3).sub.4 (232.2 mg, 0.2010
mmol), K.sub.2CO.sub.3(s) (833.3 mg, 6.029 mmol). The mixture was
sparged with Ar.sub.(g), then sealed and stirred for 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with EtOAc (200 mL), and the resulting solution
was extracted with water (2.times.50 mL) then brine (25 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by C18 reverse phase chromatography (2-75% water/ACN
with 0.1% TFA as the gradient eluent) to afford the title compound
as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH (50 mL)
and extracted with saturated NaHCO.sub.3(aq) (1.times.25 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum cleanly affording the title
compound (160.5 mg, 25% yield). MS (apci) m/z=323.1 (M+H).
Step 2: Preparation of Methyl
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)benzoate
[0527] A solution of methyl
3-(3-(2-aminopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)benzoate (from
step 1; 160.5 mg, 0.4980 mmol) in ACN (5.0 mL) was treated with NBS
(97.49 mg, 0.5478 mmol) then stirred for 16 h at ambient
temperature then concentrated under vacuum. The resulting residue
was triturated with EtOAc (10 mL) filtered to afford the title
compound (169.0 mg, 85% yield). MS (apci) m/z=403.0 [(M+H)+2],
401.0 (M+H), with Br pattern.
Step 3: Preparation of
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)benzoic
Acid
[0528] A solution of methyl
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)benzoate
(from step 2; 142.8 mg, 0.3559 mmol) in 1:1 THF:MeOH (3.6 mL) was
treated with 2 M KOH.sub.(aq) (889.8 .mu.L, 1.780 mmol) then
stirred for 1 h at 70.degree. C. After cooling to ambient
temperature, the reaction mixture was diluted with water (10 mL).
The resulting suspension was filtered, and the filter cake was
washed with water (2.times.10 mL). The solids were dried under high
vacuum for 16 h at 55.degree. C. to afford the title compound (93.8
mg, 68% yield). MS (apci) m/z=389.0 [(M+H)+2], 387.0 (M+H), with Br
pattern.
Step 4: Preparation of
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)-N-methylbenza-
mide
[0529] A solution of methyl
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)benzoic
acid (from step 3; 93.8 mg, 0.242 mmol) in DMF (2.5 mL) was treated
with methylamine hydrochloride (49.1 mg, 0.727 mmol), HATU (110 mg,
0.291 mmol), and DIPEA (211 .mu.L, 1.21 mmol). The resulting
mixture was stirred for 2 h at ambient temperature before directly
chromatographing the reaction mixture using C18 reverse phase
chromatography (5-95% water/ACN w/0.1% TFA as the gradient eluent)
to afford the title compound as the TFA salt. The TFA salt was
dissolved in 4:1 DCM:iPrOH (25 mL) and extracted with saturated
NaHCO.sub.3(aq) (1.times.10 mL). The organic extracts were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum cleanly affording the title compound (47.6 mg, 49% yield).
MS (apci) m/z=402.0 [(M+H)+2], 400.0 (M+H), with Br pattern.
[0530] Intermediate M2
##STR00149##
6-(2-amino-5-bromopyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3 (2H)-one
Step 1: Preparation of
6-(2-aminopyridin-3-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0531] A solution of t-Butyl
3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-ylcarbamate
(923.4 mg, 2.884 mmol) in 4:1 dioxane:water (15 mL) was treated
with 6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(Intermediate X8; 807.5 mg, 3.028 mmol), Pd(PPh.sub.3).sub.4(333.2
mg, 0.2884 mmol), K.sub.2CO.sub.3(s) (1195 mg, 8.652 mmol). The
mixture was sparged with Ar.sub.(g), then sealed and stirred for 16
h at 100.degree. C. After cooling to ambient temperature, the
reaction mixture was diluted with 4:1 DCM:iPrOH (100 mL), and the
resulting solution was extracted with water (2.times.50 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by silica chromatography (1-25% DCM/MeOH as the
gradient eluent) to afford the title compound (505.5 mg, 54%
yield). MS (apci) m/z=325.1 (M+H).
Step 2: Preparation of
6-(2-aminopyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3(2-
H)-one
[0532] A cold (0.degree. C.) solution of
6-(2-aminopyridin-3-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
(from step 1; 12.7 mg, 0.0392 mmol) in ACN (0.8 mL) was treated
with SO.sub.2Cl.sub.2 (6.33 .mu.L, 0.0783 mmol) then stirred for 30
min at ambient temperature. The resulting mixture was quenched with
the addition of saturated NaHCO.sub.3(aq) (10 mL). The resulting
biphasic mixture was extracted with 4:1 DCM:iPrOH (2.times.25 mL).
The combined organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
afford the title compound (13.1 mg, 85% yield). MS (apci) m/z=396.9
[(M+H)+4], 394.9 [(M+H)+2], 392.9 (M+H), with di Cl pattern.
Step 3: Preparation of
6-(2-amino-5-bromopyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3(2H)-one
[0533] A solution of
6-(2-aminopyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3(2-
H)-one (from step 2; 12.0 mg, 0.0305 mmol) in ACN (0.6 mL) was
treated with NBS (5.97 mg, 0.0336 mmol) then stirred for 16 h at
ambient temperature. The resulting mixture was diluted with 4:1
DCM:iPrOH (25 mL) and extracted with water (2.times.10 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum to afford the title compound
(14.4 mg, 100% yield). MS (apci) m/z=474.9 [(M+H)+4], 472.9
[(M+H)+2], 470.9 (M+H) with di Cl pattern.
[0534] Intermediate M3
##STR00150##
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3 (2H)-one
[0535] 3,5-Dibromopyrazin-2-amine (0.417 g, 1.65 mmol) and sodium
carbonate (2.62 ml, 5.24 mmol) were added to a solution of
2-(2,6-dichloro-3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxabo-
rolan-2-yl)pyridazin-3 (2H)-one (Intermediate R17; 0.640 g, 1.50
mmol) in 1,4-dioxane (15.0 ml, 1.50 mmol). The reaction mixture was
stirred at 55.degree. C. for 20 hrs. The reaction was quenched with
water (50 mL) and extracted with DCM. The combined organic extracts
were washed with water and brine, dried over Na.sub.2SO.sub.4 and
concentrated in vacuo. The residue was triturated with DCM (50 mL)
and filtered to obtain title compound (0.207 g, 0.438 mmol, 29.2%
yield). MS (apci) m/z=477.9 [(M+H)+4], 475.9 [(M+H)+2], 473.9
(M+H), 471.9 (M-H) with di Cl+Br pattern.
[0536] Intermediate M4
##STR00151##
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-me-
thylpyridazin-3 (2H)-one
[0537] A mixture of 3,5-dibromopyrazin-2-amine (0.585 g, 2.31
mmol),
2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methyl-6-(4,4,5,5-tetramethyl-1,3,-
2-dioxaborolan-2-yl)pyridazin-3(2H)-one (Intermediate R16; 0.927 g,
2.10 mmol), Pd(PPh.sub.3).sub.4(0.182 g, 0.158 mmol) and
Na.sub.2CO.sub.3 (2.21 mL, 4.41 mmol) in dioxane (10.5 ml, 2.10
mmol) was stirred at 55.degree. C. for 8 hours. The reaction was
quenched with water and extracted with DCM. The combined organic
extracts were washed with water and brine, dried over
Na.sub.2SO.sub.4 and concentrated in vacuo. The crude residue was
purified by flash chromatography (1-9% MeOH in DCM) to give the
title compound (0.187 g, 0.384 mmol, 18.3% yield). MS (apci)
m/z=491.9 [(M+H)+4], 489.9 [(M+H)+2], 487.9 (M+H), 485.9 (M-H) with
di Cl+Br pattern
Preparation of Synthetic Examples
Example 1
##STR00152##
[0538]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimetho-
xyphenyl)pyridazin-3 (2H)-one
[0539] A solution of
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3(2H)-one (Intermediate R8; 275.0 mg, 0.7677 mmol) in 4:1
dioxane:water (7.7 mL) was treated with
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2; 204.8 mg, 0.8061 mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2
(63.16 mg, 0.07677 mmol), K.sub.2CO.sub.3(s) (318.3 mg, 2.303
mmol). The resulting mixture was sparged with Ar.sub.(g), then
sealed and stirred for 16 h at 100.degree. C. After cooling to
ambient temperature, the reaction mixture was diluted with 4:1
DCM:iPrOH (150 mL), and extracted with water (2.times.50 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by silica chromatography (5-80% DCM/Acetone as the
gradient eluent) to afford the title compound (189.6 mg, 61%
yield). MS (apci) m/z=406.1 (M+H). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.67-8.65 (d, 1H), 8.48 (s, 1H), 8.30 (s,
1H), 8.02 (s, 1H), 7.31 (s, 2H), 7.22-7.20 (d, 1H), 6.86-6.85 (d,
2H), 6.63-6.62 (m, 1H), 3.89 (s, 3H), 3.79 (s, 6H).
[0540] The following compounds shown in Table 1 were prepared
according the method used in Example 1 using the appropriate
3-bromo-5-(pyrazoyl)pyrazin-2-amines (Intermediates L2, L14, L15,
L16, L19) and
2-(Aryl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridazi-
n-3(2H)-one (Intermediate R1, R8, R9; 1.00-1.2 equivalents).
Reaction progression in each was followed by LCMS and reaction time
was adjusted as necessary. All compounds were purified using a
method similar to that followed in Example 1 utilizing the
appropriate gradient eluent.
TABLE-US-00018 TABLE 1 MS (apci) Ex # Structure Name m/z 2*
##STR00153## 6-(3-amino-6-(1- methyl-1H-pyrazol-
4-yl)pyrazin-2-yl)-2- (3- methoxyphenyl)pyridazin- 3(2H)-one 376.0
(M + H) 3 ##STR00154## 6-(3-amino-6-(1- methyl-1H-pyrazol-
4-yl)pyrazin-2-yl)-2- (3,4- dimethoxyphenyl)pyridazin- 3(2H)-one
406.1 (M + H) 4 ##STR00155## 1-((4-(5-amino-6-(1- (3,5-
dimethoxyphenyl)-6- oxo-1,6- dihydropyridazin-3-
yl)pyrazin-2-yl)-1H- pyrazol-1- yl)methyl)cyclopropane-
1-carbonitrile 471.1 (M + H) 5 ##STR00156## 6-(3-amino-6-(1-
(4,4,4-trifluoro-2- hydroxybutyl)-1H- pyrazol-4-yl)pyrazin-
2-yl)-2-(3,5- dimethoxyphenyl)pyridazin- 3(2H)-one 518.0 (M + H) 6
##STR00157## 2-(4-(5-amino-6-(1- (3,5- dimethoxyphenyl)-6- oxo-1,6-
dihydropyridazin-3- yl)pyrazin-2-yl)-1H- pyrazol-1-yl)-2-
methylpropanenitrile 459.1 (M + H) 7 ##STR00158##
6-(3-amino-6-(1-(1- hydroxy-2- methylpropan-2-yl)- 1H-pyrazol-4-
yl)pyrazin-2-yl)-2- (3,5- dimethoxyphenyl)pyridazin- 3(2H)-one
464.1 (M + H) *EtOAc was used as the work up solvent in place of
4:1 DCM iPrOH
Example 8
##STR00159##
[0541]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3-methoxy-5-
-(trifluoromethoxy)phenyl)pyridazin-3(2H)-one
2,2,2-trifluoroacetate Salt
[0542] A suspension of
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2; 0.100 g, 0.394 mmol),
2-(3-methoxy-5-(trifluoromethoxy)phenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dio-
xaborolan-2-yl)pyridazin-3(2H)-one (Intermediate R2; 0.243 g, 0.590
mmol), Pd(PPh.sub.3).sub.4(0.0341 g, 0.0295 mmol) and 2M
Na.sub.2CO.sub.3(aq) (0.413 mL, 0.826 mmol) in dioxane (1.0 mL) was
sparged with Ar.sub.(g), then sealed and stirred for 8 h at
90.degree. C. After cooling to ambient temperature, the reaction
mixture was filtered to remove solids. The filtrate was
concentrated under vacuum, and the resulting crude residue was
purified by C18 reverse phase chromatography (5-95% ACN:water with
0.1% TFA) to afford the title compound (0.1014 g, 46% yield). MS
(apci) m/z=460.1 (M+H).
[0543] The following compounds shown in Table 2, were prepared
according the method used in Example 8 using the appropriate
3-bromo-5-(pyrazoyl)pyrazin-2-amines (Intermediates L2, L3, L5,
L17) and
2-(Aryl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridazin-3(2H)-o-
ne (Intermediate R3, R4, R6, R7, R8, R12). Reaction progression in
each was followed by LCMS and reaction time was adjusted as
necessary. All compounds were purified using a method similar to
that used in Example 8 utilizing the appropriate gradient eluent
and in each case the mono-TFA salt was isolated.
TABLE-US-00019 TABLE 2 MS (apci) Ex # Structure Name m/z 9
##STR00160## 6-(3-amino-6-(1- methyl-1H-pyrazol-4-
yl)pyrazin-2-yl)-2-(3- ethoxy-5-
(trifluoromethoxy)phenyl)pyridazin- 3(2H)-one
2,2,2-trifluoroacetate salt 474.1 (M + H) 10 ##STR00161##
6-(3-amino-6-(1- methyl-1H-pyrazol-4- yl)pyrazin-2-yl)-2-(3-
methoxy-5- (trifluoromethyl)phenyl)pyridazin- 3(2H)-one
2,2,2-trifluoroacetate salt 444.1 (M + H) 11 ##STR00162##
6-(3-amino-6-(1- methyl-1H-pyrazol-4- yl)pyrazin-2-yl)-2-(2-
chloro-3- methoxyphenyl)pyridazin- 3(2H)-one 2,2,2-
trifluoroacetate salt 410.1 (M + H) 12 ##STR00163##
6-(3-amino-6-(1- methyl-1H-pyrazol-4- yl)pyrazin-2-yl)-2-(o-
tolyl)pyridazin-3(2H)- one 2,2,2- trifluoroacetate salt 360.1 (M +
H) 13 ##STR00164## 3-(3-(3-amino-6-(1- methyl-1H-pyrazol-4-
yl)pyrazin-2-yl)-6- oxopyridazin-1(6H)- yl)-5- methoxybenzonitrile
2,2,2-trifluoroacetate salt 401.1 (M + H) 14 ##STR00165##
6-(3-amino-6-(1- methyl-1H-pyrazol-4- yl)pyrazin-2-yl)-2-(2-
chloro-3,5- dimethoxyphenyl)pyridazin- 3(2H)-one (2,2,2-
trifluoroacetate) salt 440.1 (M + H) 15 ##STR00166##
6-(3-amino-6-(1- isopropyl-1H-pyrazol- 4-yl)pyrazin-2-yl)-2- (3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 2,2,2- trifluoroacetate 434.1
(M + H) 16 ##STR00167## 6-(3-amino-6-(1- (pentan-3-yl)-1H-
pyrazol-4-yl)pyrazin-2- yl)-2-(3,5- dimethoxyphenyl)pyridazin-
3(2H)-one 2,2,2- trifluoroacetate salt 462.2 (M + H) 17
##STR00168## 6-(3-amino-6-(1- (cyclopropylmethyl)- 1H-pyrazol-4-
yl)pyrazin-2-yl)-2-(3,5- dimethoxyphenyl)pyridazin- 3(2H)-one
2,2,2- trifluoroacetate salt 446.2 (M + H) 18 ##STR00169##
6-(3-amino-6-(1-(2- hydroxy-2- methylpropyl)-1H-
pyrazol-4-yl)pyrazin-2- yl)-2-(3,5- dimethoxyphenyl)pyridazin-
3(2H)-one 2,2,2- trifluoroacetate salt 464.2 (M + H)
Example 19
##STR00170##
[0544]
6-(3-amino-6-(1-(2-morpholinoethyl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-
-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0545] A suspension of
3-bromo-5-(1-(2-morpholinoethyl)-1H-pyrazol-4-yl)pyrazin-2-amine
(Intermediate L8; 0.044 g, 0.125 mmol),
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3(2H)-one (Intermediate R8; 0.0892 g, 0.249 mmol),
Pd(PPh.sub.3).sub.4 (0.0108 g, 0.00934 mmol) and 2M
Na.sub.2CO.sub.3(aq) (0.131 mL, 0.262 mmol) in dioxane (1 mL) was
sparged with Ar.sub.(g), then sealed and stirred for 8 h at
90.degree. C. After cooling to ambient temperature, the reaction
mixture was filtered to remove solids. The filtrate was
concentrated under vacuum, and the resulting crude residue was
purified by silica chromatography (5-95% DCM:(DCM:MeOH:NHOH.sub.4
(90:10:1)) to afford the title compound (0.0538 g, 86% yield). MS
(apci) m/z=505.2 (M+H).
[0546] The following compounds shown in Table 3 were prepared
according the method used in Example 19 using the appropriate
3-bromo-5-(pyrazoyl)pyrazin-2-amines (Intermediates L2, L4, L7) and
2-(Aryl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridazin-3(2H)-o-
ne (Intermediate R5, R8). Reaction progression in each was followed
by LCMS and reaction time was adjusted as necessary. All compounds
were purified using a method similar to that followed in Example 19
utilizing the appropriate gradient eluent.
TABLE-US-00020 TABLE 3 MS (apci) Ex # Structure Name m/z 20
##STR00171## 6-(3-amino-6-(1- methyl-1H-pyrazol-4-
yl)pyrazin-2-yl)-2-(3- isopropoxy-5- methoxyphenyl)pyridazin-
3(2H)-one 434.2 (M + H) 21 ##STR00172## 6-(3-amino-6-(1-
isobutyl-1H-pyrazol- 4-yl)pyrazin-2-yl)-2- (3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 448.2 (M + H) 22 ##STR00173##
6-(3-amino-6-(1- cyclobutyl-1H- pyrazol-4-yl)pyrazin- 2-yl)-2-(3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 446.2 (M + H)
Example 23
##STR00174##
[0547]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,6-dichlor-
o-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0548] A cold (0.degree. C.) solution of
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphen-
yl)pyridazin-3(2H)-one (Example 1; 13.6 mg, 0.0335 mmol) in ACN
(0.7 mL) was treated with SO.sub.2Cl.sub.2 (5.42 .mu.L, 0.0671
mmol) then stirred for 30 min at ambient temperature. The resulting
mixture was quenched with the addition of saturated NaHCO.sub.3(aq)
(10 mL). The resulting biphasic mixture was extracted with 4:1
DCM:iPrOH (2.times.25 mL). The combined organic extracts were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The resulting crude residue was purified by silica
chromatography (5-50% DCM/Acetone as the gradient eluent) to afford
the title compound (189.6 mg, 61% yield). MS (apci) m/z=476.0
[(M+H)+2], 474.1 (M+H), with di Cl pattern. .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.73-8.70 (d, 1H), 8.47 (s, 1H), 8.27 (s,
1H), 8.00 (s, 1H), 7.30-7.27 (d, 1H), 7.09 (s, 1H), 7.03 (s, 2H),
3.94 (s, 6H), 3.85 (s, 3H).
Example 24
##STR00175##
[0549]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-bromo-3,5-
-dimethoxyphenyl)pyridazin-3(2H)-one 2,2,2-trifluoroacetate
Salt
[0550] A cold (0.degree. C.) solution of
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphen-
yl)pyridazin-3(2H)-one (Example 1; 100 mg, 0.247 mmol) in DCM (1.23
mL) was treated with NBS (43.9 mg, 0.247 mmol) then stirred for 3 h
at ambient temperature. The resulting mixture was diluted with
Ethyl Acetate, washed with Brine (2x), then dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting crude residue was purified by silica gel chromatography
(30-100% EtOAc/Hexanes as the gradient eluent) to afford the title
compound in about 75% purity (17.4 mg, 15% yield). Additional
purification by C18 reverse phase chromatography (5-95%
Acetonitrile in Water with 0.1% TFA as gradient eluent) provided
clean title compound. MS (apci) m/z=484.0 (M+H), with di Br
pattern. 1H NMR (400 MHz, CDCl.sub.3) .delta. 8.67-8.64 (d, 1H),
8.26 (s, 1H), 7.92 (s, 1H), 7.83 (s, 1H), 7.20-7.17 (d, 1H),
6.65-6.60 (m, 2H), 6.27 (broad s, 2H), 3.96 (s, 3H), 3.92 (s, 3H),
3.82 (s, 3H).
Example 25
##STR00176##
[0551]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,6-dibromo-
-3,5-dimethoxyphenyl)pyridazin-3(2H)-one 2,2,2-trifluoroacetate
Salt
[0552] A cold (0.degree. C.) solution of
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphen-
yl)pyridazin-3(2H)-one (Example 1; 20 mg, 0.0493 mmol) in DCM
(0.247 mL mL) was treated with NBS (15 mg, 0.0843 mmol) then
stirred overnight at ambient temperature. The resulting mixture was
diluted with Ethyl Acetate, washed with Brine (2x), then dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The resulting crude residue was purified by C18 reverse
phase chromatography (5-95% Acetonitrile in Water with 0.1% TFA as
the gradient) to afford the title compound (7 mg, 25% yield). MS
(apci) m/z=564.0 (M+H), with di Br pattern. .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 8.84-8.81 (d, 1H), 8.36 (s, 1H), 8.16 (s, 1H),
8.03 (s, 1H), 7.27-7.24 (d, 1H), 6.95 (s, 1H), 4.00 (s, 6H), 3.94
(s, 3H).
Example 26
##STR00177##
[0553]
(R)-6-(3-amino-6-(1-((5,5-dimethylmorpholin-2-yl)methyl)-1H-pyrazol-
-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of tert-butyl
(R)-2-((4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimethylmorpholine-4-carbox-
ylate
[0554] A suspension of tert-butyl
(R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimet-
hylmorpholine-4-carboxylate (Intermediate L12; 100 mg, 0.214 mmol),
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3(2H)-one (Intermediate R8; 192 mg, 0.535 mmol),
Pd(PPh.sub.3).sub.4(24.7 mg, 0.0214 mmol), and 2M
Na.sub.2CO.sub.3(aq) (321 .mu.L, 0.642 mmol) in dioxane (2.14 mL)
was sparged with Ar.sub.(g), then sealed and stirred 2 h at
90.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with EtOAc and filtered to remove solids. The
filtrate was concentrated under vacuum, and the resulting crude
residue was purified by silica chromatography (60-100% EtOAc in
Hexanes) to afford the title compound (35 mg, 26% yield). MS (apci)
m/z=619.2 (M+H).
Step 2: Preparation of
(R)-6-(3-amino-6-(1-((5,5-dimethylmorpholin-2-yl)methyl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0555] The tert-butyl
(R)-2-((4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimethylmorpholine-4-carbox-
ylate (35 mg, 0.057 mmol) was dissolved in 1:1 TFA:DCM (8.0 mL)
then stirred 1 h at ambient temperature. The reaction mixture was
concentrated under vacuum then purified by C18 reverse phase
chromatography (5-95% ACN in H.sub.2O with 0.1% TFA as the gradient
as the gradient eluent). The chromatographic fractions containing
the title compound were combined then neutralized with saturated
NaHCO.sub.3(aq) and extracted with DCM. The organic extracts were
then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum to afford the title compound (24.5 mg,
84% yield). MS (apci) m/z=519.2 (M+H).
Example 27
##STR00178##
[0556]
(S)-6-(3-amino-6-(1-((5,5-dimethylmorpholin-2-yl)methyl)-1H-pyrazol-
-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of Tert-Butyl
(S)-2-((4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimethylmorpholine-4-carbox-
ylate
[0557] The title compound was prepared and isolated according to
the method described in step 1 of Example 26, using tert-butyl
(S)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimet-
hylmorpholine-4-carboxylate (Intermediate L11) in place of
tert-butyl
(R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimet-
hylmorpholine-4-carboxylate (Intermediate L12). The title compound
was carried on to step 2, without silica chromatography. MS (apci)
m/z=446.2 (M+H).
Step 2: Preparation of
(S)-6-(3-amino-6-(1-((5,5-dimethylmorpholin-2-yl)methyl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0558] The title compound was prepared and isolated according to
the method described in step 2 of Example 26, using the crude
residue from step 1 and neutralizing with 1 M NaOH.sub.(aq) instead
of NaHCO.sub.3(aq) resulting in a 46% yield (39 mg). MS (apci)
m/z=519.2 (M+H).
Example 28
##STR00179##
[0559]
6-(3-amino-6-(1-(((2S,5R)-5-methylmorpholin-2-yl)methyl)-1H-pyrazol-
-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of tert-butyl
(2S,5R)-2-((4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5-methylmorpholine-4-carbox-
ylate
[0560] The title compound was prepared and isolated according to
the method described in step 1 of Example 26, with the exception
that the reaction required overnight stirring at 90.degree. C. and
utilized tert-butyl
(2S,5R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5-met-
hylmorpholine-4-carboxylate (Intermediate L10) in place of
tert-butyl
(R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimet-
hylmorpholine-4-carboxylate (Intermediate L12). The title compound
was carried on to step 2, without silica chromatography (23 mg, 34%
yield). MS (apci) m/z=605.2 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(((2S,5R)-5-methylmorpholin-2-yl)methyl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0561] Using the crude residue from step 1, the title compound was
prepared, isolated, purified and neutralized according to the
method described in step 2 of Example 26, with the exception that
the reaction was allowed to stir 2 d at ambient temperature prior
to isolation/purification (7.0 mg, 84% yield). MS (apci) m/z=505.2
(M+H).
Example 29
##STR00180##
[0562]
(R)-6-(6-(1-((7-oxa-4-azaspiro[2.5]octan-6-yl)methyl)-1H-pyrazol-4--
yl)-3-aminopyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3
(2H)-one
Step 1: Preparation of Tert-Butyl
(R)-6-((4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-7-oxa-4-azaspiro[2.5]octane-4-c-
arboxylate
[0563] The title compound was prepared and isolated according to
the method described in step 1 of Example 26, using of tert-butyl
(R)-6-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-7-oxa-4-a-
zaspiro[2.5]octane-4-carboxylate (Intermediate L13) in place of
tert-butyl
(R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimet-
hylmorpholine-4-carboxylate (Intermediate L12). The title compound
was purified by silica chromatography (30-75% EtOAc in Hexanes)
which provide the title compound in 38% yield (15 mg). MS (apci)
m/z=617.2 (M+H).
Step 2: Preparation of
(R)-6-(6-(1-((7-oxa-4-azaspiro[2.5]octan-6-yl)methyl)-1H-pyrazol-4-yl)-3--
aminopyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0564] Using the purified material from step 1, the title compound
was prepared, isolated, purified and neutralized according to the
method described in step 2 of Example 27 (12.1 mg, 96% yield). MS
(apci) m/z=517.2 (M+H).
Example 30
##STR00181##
[0565]
6-(3-amino-6-(1-(((2R,5R)-5-methylmorpholin-2-yl)methy)-1H-pyrazol--
4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of tert-butyl
(2R,5R)-2-((4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5-methylmorpholine-4-carbox-
ylate
[0566] The title compound was prepared and isolated according to
the method described in step 1 of Example 26, using of tert-butyl
(2R,5R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5-met-
hylmorpholine-4-carboxylate (Intermediate L9) in place of
tert-butyl
(R)-2-((4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)methyl)-5,5-dimet-
hylmorpholine-4-carboxylate (Intermediate L12). The title compound
was purified by silica chromatography (30-75% EtOAc in Hexanes)
which provided the title compound in 31% yield (23 mg). MS (apci)
m/z=605.2 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(((2R,5R)-5-methylmorpholin-2-yl)methyl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0567] Using the purified material from step 1, the title compound
was prepared, isolated, purified according to the method described
in step 2 of Example 26 which provided the title compound in 91%
yield (17.4 mg). MS (apci) m/z=505.2 (M+H).
Example 31
##STR00182##
[0568]
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3-
,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of Tert-Butyl
4-(4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin-3-yl)-
pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0569] A suspension of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 398.9 mg, 0.9423 mmol),
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3(2H)-one (Intermediate R8; 371.3 mg, 1.037 mmol),
PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (77.52 mg, 0.09423 mmol) and
K.sub.2CO.sub.3(s) (390.7 mg, 2.827 mmol) in 4:1 dioxane:water (10
mL) was sparged with Ar.sub.(g), then sealed and stirred 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (200 mL) and washed with
water (2.times.50 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The crude residue was purified by silica chromatography
(5-95% DCM-Acetone as the gradient eluent) to afford the title
compound which was immediately carried on to step 2. MS (apci)
m/z=575.3 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one
[0570] The tert-butyl
4-(4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin-3-yl)-
pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate was
dissolved in 1:1 TFA:DCM (5.0 mL) then stirred 30 min at ambient
temperature. The reaction mixture was concentrated under vacuum
then purified by C18 reverse phase chromatography (5-95% ACN in
H.sub.2O with 0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (213.2 mg, 48% yield). MS (apci)
m/z=475.2 (M+H).
Example 32
##STR00183##
[0571]
6-(3-amino-6-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2--
yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0572] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 31; 187.2 mg, 0.3945
mmol) in 1:1 DCM:MeOH (4 mL) was treated with formaldehyde (592.9
.mu.L, 7.890 mmol) and stirred 15 min at ambient temperature. The
resulting reaction mixture was treated with sodium
triacetoxyborohydride (334.4 mg, 1.578 mmol) and stirred 16 h at
ambient temperature. The reaction mixture was concentrated under
vacuum then purified by C18 reverse phase chromatography (5-95%
water-ACN w/0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
afford the title compound along with imine contaminants. The
residue then was dissolved in 1:1:1 TFA:ACN:water (6 mL) and
stirred for 15 min at ambient temperature then concentrated under
vacuum. The resultant residue was dissolved in 4:1 DCM:iPrOH (50
mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25 mL).
The organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The residue was purified
further by silica chromatography (1-30% DCM-MeOH w/2% NH.sub.4OH as
the gradient eluent) to cleanly afford the title compound (28.5 mg,
15% yield). MS (apci) m/z=489.1 (M+H). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.71-8.69 (d, 1H), 8.49 (s, 1H), 8.42 (s,
1H), 8.04 (s, 1H), 7.31 (s, 2H), 7.21-7.19 (d, 1H), 6.86-6.85 (d,
2H), 6.63-6.62 (t, 1H), 4.19-4.13 (m, 1H), 3.79 (s, 6H), 2.93-2.88
(m, 2H), 2.25 (s, 3H), 2.15-1.97 (m, 6H).
Example 33
##STR00184##
[0573]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0574] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 31; 10.6 mg, 0.0223 mmol)
in DMF (0.5 mL) was treated with 1-bromo-2-methoxyethane (2.5
.mu.L, 0.027 mmol) and K.sub.2CO.sub.3(s) (6.1 mg, 0.0447 mmol).
The resulting mixture was stirred 16 h at ambient temperature, then
additional 1-bromo-2-methoxyethane (2.5 .mu.L, 0.027 mmol) and
K.sub.2CO.sub.3(s) (6.1 mg, 0.0447 mmol) were added. The reaction
was stirred for an additional period of 24 h at ambient temperature
and subsequently diluted with EtOAc (25 mL). The EtOAc solution was
washed with water (2.times.10 mL) and brine (1.times.10 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The crude residue was
purified by silica chromatography (1-30% DCM-MeOH w/2% NH.sub.4OH
as the gradient eluent) to afford the title compound (6.1 mg, 51%
yield). MS (apci) m/z=533.2 (M+H). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.72-8.70 (d, 1H), 8.49 (s, 1H), 8.43 (s,
1H), 8.04 (s, 1H), 7.31 (s, 2H), 7.21-7.18 (d, 1H), 6.86-6.85 (d,
2H), 6.63-6.62 (t, 1H), 4.19-4.12 (m, 1H), 3.79 (s, 6H), 3.47-3.44
(t, 2H), 3.25 (s, 3H), 3.01-2.97 (m, 2H), 2.54-2.51 (m, 2H),
2.19-2.13 (m, 2H), 2.06-1.94 (m, 4H).
Example 34
##STR00185##
[0575]
6-(3-amino-6-(1-(1-(2-(trifluoromethoxy)ethyl)piperidin-4-yl)-1H-py-
razol-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
2,2,2-trifluoroacetate Salt
[0576] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 31; 100 mg, 0.211 mmol)
in DMSO (843 .mu.L) was treated with
1-bromo-2-(trifluoromethoxy)ethane (36.4 .mu.L, 0.316 mmol) and
Cs.sub.2CO.sub.3(s) (172 mg, 0.527 mmol). The resulting mixture was
stirred overnight at ambient temperature then diluted with EtOAc
and washed with water (2x) and brine. The organic extracts were
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum. The crude residue was purified by silica
chromatography (50-100% EtOAc in Hexanes as the gradient eluent).
The material was further purified by C18 reverse phase
chromatography (5-95% Acetonitrile in Water with 0.1% TFA as the
gradient eluent) to afford the title compound (15 mg, 12% yield).
MS (apci) m/z=587.2 (M+H).
Example 35
##STR00186##
[0577]
6-(3-amino-6-(1-(1-(2,2-difluoroethyl)piperidin-4-yl)-1H-pyrazol-4--
yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
2,2,2-trifluoroacetate Salt
[0578] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 31; 100 mg, 0.211 mmol)
in DMF (1.05 mL) was treated with DIPEA (110 .mu.L, 0.632 mmol) and
2,2-difluoroethyl trifluoromethanesulfonate (67.7 mg, 0.316 mmol).
The resulting mixture was stirred 4 h at ambient temperature then
diluted with EtOAc and washed with water and brine. The organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated under vacuum. The crude residue was purified by
silica chromatography (stepwise gradient eluent of 70-100% EtOAc in
Hexanes then 0-5% MeOH in EtOAc). The material was further purified
by C18 reverse phase chromatography (5-95% Acetonitrile in Water
with 0.1% TFA as the gradient eluent) to afford the title compound
(11 mg, 10% yield). MS (apci) m/z=539.2 (M+H).
Example 36
##STR00187##
[0579]
6-(3-amino-6-(1-(1-(2,2,2-trifluoroethyl)piperidin-4-yl)-1H-pyrazol-
-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0580] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 31; 100 mg, 0.211 mmol)
in DMF (1.05 mL) was treated with DIPEA (110 .mu.L, 0.632 mmol) and
2,2,2-trifluoroethyl trifluoromethanesulfonate (73.4 mg, 0.316
mmol). The resulting mixture was stirred over the weekend at
ambient temperature then diluted with a mixture of
EtOAc/water/brine. The organic extracts were washed with water and
brine then dried over anhydrous MgSO.sub.4(s), filtered and
concentrated under vacuum. The crude residue was purified by silica
chromatography (50-80% EtOAc in Hexanes as the gradient eluent) to
afford the title compound (30.2 mg, 26% yield). MS (apci) m/z=557.2
(M+H).
Example 37
##STR00188##
[0581]
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(5-
-methoxy-2-methylphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of tert-butyl
4-(4-(5-amino-6-(1-(5-methoxy-2-methylphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0582] A suspension of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 198.5 mg, 0.4689 mmol),
2-(5-methoxy-2-methylphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2--
yl)pyridazin-3(2H)-one (Intermediate R.sup.10; 320.9 mg, 0.9379
mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (38.58 mg, 0.04689 mmol)
and K.sub.2CO.sub.3(s) (194.4 mg, 1.407 mmol) in 4:1 dioxane:water
(5.0 mL) was sparged with Ar.sub.(g), then sealed and stirred 16 h
at 100.degree. C. After cooling to ambient temperature, the
reaction mixture was diluted with 4:1 DCM:iPrOH (50 mL) and washed
with water (2.times.25 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The crude residue was purified by silica chromatography
(5-60% DCM-Acetone as the gradient eluent) to afford the title
compound which was immediately carried on to step 2. MS (apci)
m/z=575.3 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(5-metho-
xy-2-methylphenyl)pyridazin-3(2H)-one
[0583] The tert-butyl
4-(4-(5-amino-6-(1-(5-methoxy-2-methylphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate was
dissolved in 1:1 TFA:DCM (5.0 mL) then stirred 1 h at ambient
temperature. The reaction mixture was concentrated under vacuum
then purified by C18 reverse phase chromatography (5-95% ACN in
H.sub.2O with 0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (102.0 mg, 47% yield). MS (apci)
m/z=459.1 (M+H).
Example 38
##STR00189##
[0584]
6-(3-amino-6-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2--
yl)-2-(5-methoxy-2-methylphenyl)pyridazin-3 (2H)-one
[0585] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(5-metho-
xy-2-methylphenyl)pyridazin-3(2H)-one (Example 37; 187.2 mg, 0.3945
mmol) in 1:1 DCM:MeOH (1.5 mL) was treated with formaldehyde (227
.mu.L, 3.02 mmol) and sodium triacetoxyborohydride (128 mg, 0.604
mmol) and stirred 16 h at ambient temperature. The reaction mixture
was concentrated under vacuum then purified by C18 reverse phase
chromatography (5-95% water-ACN w/0.1% TFA as the gradient eluent).
The chromatographic fractions containing the title compound were
dissolved in 1:1:1 TFA:ACN:water (6 mL) and stirred for 1 h at
ambient temperature then concentrated under vacuum. The residue was
diluted with 4:1 DCM:iPrOH (50 mL) and washed with saturated
NaHCO.sub.3(aq) (1.times.25 mL). The organic extracts were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to cleanly afford the title compound (55.0 mg, 77% yield).
MS (apci) m/z=473.1 (M+H).
Example 39
##STR00190##
[0586]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(5-methoxy-2-methylphenyl)pyridazin-3(2H)-one
2,2,2-trifluoroacetate Salt
[0587] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(5-metho-
xy-2-methylphenyl)pyridazin-3(2H)-one (Example 37; 22.46 mg,
0.04898 mmol) in DMSO (0.1959 mL) was treated with
1-bromo-2-methoxyethane (10.21 mg, 0.07348 mmol) and
K.sub.2CO.sub.3(s) (27.08 mg, 0.19598 mmol). The resulting mixture
was stirred overnight at 50.degree. C., then cooled to ambient
temperature. The reaction was diluted with water and extracted with
EtOAc. The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
crude residue was purified by C18 reverse phase chromatography
(5-95% Acetonitrile in Water with 0.1% TFA as the gradient eluent)
to afford the title compound (6.5 mg, 26% yield). MS (apci)
m/z=517.3 (M+H).
Example 40
##STR00191##
[0588]
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-
-fluoro-5-methoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of Tert-Butyl
4-(4-(5-amino-6-(1-(2-fluoro-5-methoxyphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0589] A suspension of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 48.06 mg, 0.1135 mmol),
2-(2-fluoro-5-methoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2--
yl)pyridazin-3(2H)-one (Intermediate R.sup.14; 117.9 mg, 0.1703
mmol), Pd(PPh.sub.3).sub.4(13.12 mg, 0.01135 mmol), and 2M
Na.sub.2CO.sub.3(aq) (170.3 .mu.L, 0.3406 mmol) in dioxane (1135
.mu.L) was sparged with Ar.sub.(g), then sealed and stirred
overnight at 90.degree. C. After cooling to ambient temperature,
the reaction mixture was partitioned between EtOAc and water then
the aqueous extracts were washed with EtOAc. The combined organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated under vacuum. The resulting crude residue was
purified by silica chromatography (0-15% MeOH in EtOAc as the
gradient eluent) to afford the title compound (15 mg, 23% yield).
MS (apci) m/z=563.3 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-fluor-
o-5-methoxyphenyl)pyridazin-3(2H)-one
[0590] The tert-butyl
4-(4-(5-amino-6-(1-(2-fluoro-5-methoxyphenyl)-6-oxo-1,6-dihydropyridazin--
3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate (15 mg,
0.027 mmol) was dissolved in 1:1 TFA:DCM (2.0 mL) then stirred
overnight at ambient temperature. The reaction mixture was
concentrated under vacuum then purified by C18 reverse phase
chromatography (5-95% ACN in H.sub.2O with 0.1% TFA as the gradient
eluent). The chromatographic fractions containing the title
compound were combined then neutralized with saturated
NaHCO.sub.3(aq) and extracted with DCM (3x). The organic extracts
were then dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum to afford the title compound (6.2 mg, 50%
yield). MS (apci) m/z=463.2 (M+H).
Example 41
##STR00192##
[0591]
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-
-fluoro-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
Step 1: Preparation of tert-butyl
4-(4-(5-amino-6-(1-(2-fluoro-3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0592] A suspension of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 69.88 mg, 0.1651 mmol),
2-(2-fluoro-3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3(2H)-one (Intermediate R13; 124.2 mg, 0.2476
mmol), Pd(PPh.sub.3).sub.4(19.08 mg, 0.01651 mmol), and 2M
Na.sub.2CO.sub.3(aq) (247.6 .mu.L, 0.4952 mmol) in dioxane (1651
.mu.L) was sparged with Ar.sub.(g), then sealed and stirred
overnight at 90.degree. C. After cooling to ambient temperature,
the reaction mixture was partitioned between EtOAc and water then
the aqueous extracts were washed with EtOAc. The combined organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated under vacuum. The resulting crude residue was
purified by silica chromatography (0-15% MeOH in EtOAc as the
gradient eluent) to afford the title compound (50 mg, 51% yield).
MS (apci) m/z=593.3 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-fluor-
o-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0593] The tert-butyl
4-(4-(5-amino-6-(1-(2-fluoro-3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate (50
mg, 0.084 mmol) was dissolved in 1:1 TFA:DCM (2.0 mL) then stirred
overnight at ambient temperature. The reaction mixture was
concentrated under vacuum then purified by C18 reverse phase
chromatography (5-95% ACN in H.sub.2O as the gradient eluent). The
chromatographic fractions containing the title compound were
combined then neutralized with saturated NaHCO.sub.3(aq) and
extracted with DCM (3x). The organic extracts were then dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to afford the title compound (37.0 mg, 89% yield). MS (apci)
m/z=493.2 (M+H).
Example 42
##STR00193##
[0594]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(2-fluoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
2,2,2-trifluoroacetate Salt
[0595] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-fluor-
o-3,5-dimethoxyphenyl)pyridazin-3(2H)-one (Example 41; 10 mg, 0.020
mmol) in DMSO (81 .mu.L) was treated with 1-bromo-2-methoxyethane
(6.91 .mu.L, 0.0734762 mmol) and K.sub.2CO.sub.3(s) (11 mg, 0.081
mmol). The resulting mixture was stirred 3 h at ambient
temperature. The reaction was diluted with water and extracted with
DCM. The organic extracts were washed with brine and were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The crude residue was purified by C18 reverse phase
chromatography (25-75% ACN in water with 0.1% TFA as the gradient
eluent) to afford the title compound (4 mg, 36% yield). MS (apci)
m/z=551.2 (M+H).
Example 43
##STR00194##
[0596]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimetho-
xyphenyl)-4-methylpyridazin-3 (2H)-one
[0597] A solution of
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2; 101.7 mg, 0.4003 mmol) in 4:1 dioxane:water (4.0 mL) was
treated with
2-(3,5-dimethoxyphenyl)-4-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3(2H)-one (Intermediate R18; 260.724 mg, 0.7004
mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (32.93 mg, 0.04003 mmol),
K.sub.2CO.sub.3(s) (166.0 mg, 1.201 mmol). The resulting mixture
was sparged with Ar.sub.(g), then sealed and stirred for 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (50 mL), and extracted with
water (2.times.25 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The resulting crude residue was purified by silica
chromatography (5-75% DCM/Acetone as the gradient eluent). The
chromatographic fractions containing the title compound were
combined, concentrated under vacuum then triturated with DCM (20
mL). The resulting suspension was filtered and the solids collected
were dried under vacuum to cleanly afford the title compound (90.8
mg, 54% yield). MS (apci) m/z=420.1 (M+H). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.55-8.54 (d, 1H), 8.46 (s, 1H), 8.32 (s,
1H), 8.07 (s, 1H), 7.30 (s, 2H), 6.84-6.83 (d, 2H), 6.62-6.61 (t,
1H), 3.90 (s, 3H), 3.79 (s, 6H), 2.29 (s, 3H).
Example 44
##STR00195##
[0598]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimetho-
xyphenyl)-4-ethylpyridazin-3(2H)-one 2,2,2-trifluoroacetate
salt
[0599] A suspension of
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2; 0.025 g, 0.098 mmol),
2-(3,5-dimethoxyphenyl)-4-ethyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-
-2-yl)pyridazin-3(2H)-one (Intermediate R26; 0.042 g, 0.11 mmol),
Pd(PPh.sub.3).sub.4 (0.0085 g, 0.0074 mmol) and 2M
Na.sub.2CO.sub.3(aq) (0.10 mL, 0.21 mmol) in dioxane (1.0 mL) was
sparged with Ar.sub.(g), then sealed and stirred for 8 h at
90.degree. C. After cooling to ambient temperature, the reaction
mixture was filtered to remove solids. The filtrate was
concentrated under vacuum, and the resulting crude residue was
purified by C18 reverse phase chromatography (5-95% ACN:water with
0.1% TFA) to afford the title compound as the TFA salt (0.0056 g,
11% yield). MS (apci) m/z=434.1 (M+H).
[0600] The following compounds shown in Table 4 were prepared
according the method used for the synthesis of Example 44 using
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2) and
2-(3,5-dimethoxyphenyl)-4-alkyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-
-2-yl)pyridazin-3(2H)-one (Intermediates R25, R27). Reaction
progression in each was followed by LCMS and reaction time was
adjusted as necessary. All compounds were purified using a method
similar to that used in Example 44 utilizing the appropriate
gradient eluent.
TABLE-US-00021 TABLE 4 MS (apci) Ex # Structure Name m/z 45
##STR00196## 6-(3-amino-6-(1- methyl-1H-pyrazol-
4-yl)pyrazin-2-yl)-2- (3,5- dimethoxyphenyl)-4- propylpyridazin-
3(2H)-one 2,2,2- trifluoroacetate salt 488.2 (M + H) 46
##STR00197## 6-(3-amino-6-(1- methyl-1H-pyrazol-
4-yl)pyrazin-2-yl)-2- (3,5- dimethoxyphenyl)-4- isobutylpyridazin-
3(2H)-one 2,2,2- trifluoroacetate salt 462.2 (M + H)
Example 47
##STR00198##
[0601]
6-(3-amino-6-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dimetho-
xyphenyl)-4-isopropylpyridazin-3 (2H)-one
[0602] A solution of
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2; 100.0 mg, 0.3936 mmol) in 4:1 dioxane:water (4.0 mL) was
treated with
2-(3,5-dimethoxyphenyl)-4-isopropyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxabor-
olan-2-yl)pyridazin-3(2H)-one (Intermediate R23; 173.3 mg, 0.4329
mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (32.93 mg, 0.04003 mmol),
K.sub.2CO.sub.3(s) (163.2 mg, 1.181 mmol). The resulting mixture
was sparged with Ar.sub.(g), then sealed and stirred for 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (50 mL), and extracted with
water (2.times.25 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The resulting crude residue was purified by silica
chromatography (5-60% DCM/Acetone as the gradient eluent) to
cleanly afford the title compound (27.1 mg, 15% yield). MS (apci)
m/z=448.1 (M+H). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 8.47
(s, 1H), 8.40 (s, 1H), 8.26 (s, 1H), 8.01 (s, 1H), 7.30 (s, 2H),
6.85-6.84 (d, 2H), 6.62-6.61 (t, 1H), 3.91 (s, 3H), 3.79 (s, 6H),
3.24-3.17 (m, 1H), 1.30-1.29 (d, 6H).
[0603] The following compounds shown in Table 5 were prepared
according the method used for the synthesis of Example 47 using
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L2) and
2-(3,5-dimethoxyphenyl)-4-alkyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-
-2-yl)pyridazin-3(2H)-one (Intermediates R22, R24). Reaction
progression in each was followed by LCMS and reaction time was
adjusted as necessary. All compounds were purified using a method
similar to that used in Example 47 utilizing the appropriate
gradient eluent.
TABLE-US-00022 TABLE 5 MS (apci) Ex # Structure Name m/z 48
##STR00199## 6-(3-amino-6-(1-methyl- 1H-pyrazol-4-yl)pyrazin-
2-yl)-4-cyclopropyl-2- (3,5- dimethoxyphenyl)pyridazin- 3(2H)-one
446.1 (M + H) 49 ##STR00200## 6-(3-amino-6-(1-methyl-
1H-pyrazol-4-yl)pyrazin- 2-yl)-4-cyclobutyl-2-(3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 460.1 (M + H)
Example 50
##STR00201##
[0604]
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3-
,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one dihydrochloride
salt
Step 1: Preparation of tert-butyl
4-(4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0605] A mixture of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 398 mg, 0.940 mmol),
2-(3,5-dimethoxyphenyl)-4-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3 (2H)-one (Intermediate R18; 350 mg, 0.940 mmol),
Pd(PPh.sub.3).sub.4(109 mg, 0.0940 mmol), 2M Na.sub.2CO.sub.3(aq)
(1.41 mL, 0.940 mmol) was suspended in dioxane (1.88 mL). The
resulting mixture was purged with N.sub.2(g) for 6 min, then was
sealed and stirred for 16 h at 90.degree. C. After cooling to
ambient temperature, the reaction mixture was diluted with DCM (50
mL) and washed with water (2.times.15 mL). The organic extracts
were dried over anhydrous MgSO.sub.4(s), filtered and concentrated
under vacuum. The crude residue was purified by silica
chromatography (1-55% acetone/hexanes as the gradient eluent) to
afford the title compound (425 mg, 77% yield). MS (apci) m/z=589.3
(M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)-4-methylpyridazin-3(2H)-one Dihydrochloride Salt
[0606] The tert-butyl
4-(4-(5-amino-6-(1-(3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
(420 mg, 0.713 mmol) was dissolved in TFA (2.0 mL) then stirred 2 h
at ambient temperature. The reaction mixture was concentrated under
vacuum then dissolved in DCM (2 mL) and treated with 2 N HCl in
dioxane (5 mL). The resulting suspension was filtered and the
solids were rinsed with ACN and dried under vacuum to afford the
title compound (325 mg, 81% yield). MS (apci) m/z=489.2 (M+H).
.sup.1H NMR (400 MHz, MeOH-d.sub.4) .delta. 8.59 (d, 1H), 8.44 (d,
1H), 8.31 (s, 1H), 8.14 (s, 1H), 6.80 (s, 1H), 6.79 (s, 1H), 6.61
(t, 1H), 4.68-4.61 (m, 1H), 3.82 (s, 6H), 3.6-3.56 (m, 2H),
3.25-3.2 (m, 2H), 2.3 (s, 3H), 2.35-2.32 (m, 4H).
Example 51
##STR00202##
[0607]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)-4-methylpyridazin-3
(2H)-one
[0608] A solution of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3,5-dim-
ethoxyphenyl)-4-methylpyridazin-3(2H)-one dihydrochloride salt
(Example 50; 50 mg, 0.089 mmol) in DMF (1.8 mL) was treated with
1-bromo-2-methoxyethane (10 .mu.L, 0.11 mmol) and
K.sub.2CO.sub.3(s) (49 mg, 0.36 mmol) under an atmosphere of
N.sub.2(g). The resulting mixture was stirred overnight at ambient
temperature, and then additional 1-bromo-2-methoxyethane (20 .mu.L,
0.21 mmol) and K.sub.2CO.sub.3(s) (98 mg, 0.71 mmol) and the
reaction was stirred for an additional period of 48 h at ambient
temperature. The reaction mixture was diluted with 5% iPrOH/DCM (50
mL) and washed with water (10 mL) diluted. The organic extracts
were dried over anhydrous MgSO.sub.4(s), filtered and concentrated
under vacuum. The crude residue was purified by silica
chromatography (2-10% MeOH/DCM as the gradient eluent) to afford
the title compound (33 mg, 68% yield). MS (apci) m/z=547.3 (M+H).
.sup.1H NMR (400 MHz, CDCl3-d) .delta. 8.44 (d, 1H), 8.25 (s, 1H),
7.91 (d, 2H), 6.79 (d, 2H), 6.51 (t, 1H), 6.48-6.3 (brs, 2H),
4.26-4.18 (m, 1H), 3.81 (s, 6H), 3.53 (t, 2H), 3.3 (s, 3H),
3.15-3.09 (m, 2H), 2.63 (d, 2H), 2.3 (s, 3H), 2.16-2.1 (m, 2H),
1.61-1.57 (m, 2H).
Example 52
##STR00203##
[0609]
3-(3-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-5-
-methyl-6-oxopyridazin-1(6H)-yl)-N,4-dimethylbenzamide
Dihydrochloride Salt
Step 1: Preparation of tert-butyl
4-(4-(5-amino-6-(1-(5-(methoxycarbonyl)-2-methylphenyl)-5-methyl-6-oxo-1,-
6-dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxy-
late
[0610] A mixture of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 505 mg, 1.19 mmol), methyl
4-methyl-3-(5-methyl-6-oxo-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl-
)pyridazin-1(6H)-yl)benzoate (Intermediate R19; 550 mg, 1.43 mmol),
Pd(PPh.sub.3).sub.4(107 mg, 0.119 mmol), 2M Na.sub.2CO.sub.3(aq)
(1.79 mL, 3.58 mmol) was suspended in dioxane (2.39 mL). The
resulting mixture was purged with N.sub.2(g) for 6 min, then was
sealed and stirred overnight at 90.degree. C. After cooling to
ambient temperature, the reaction mixture was diluted with 5%
iPrOH/DCM (100 mL) and washed with water (2.times.20 mL) and brine
(20 mL). The organic extracts were dried over anhydrous
MgSO.sub.4(s), filtered and concentrated under vacuum. The crude
residue was purified by silica chromatography (with 2-60%
acetone/DCM as the gradient eluent) to afford the title compound
(782.5 mg, 98% yield). MS (apci) m/z=601.3 (M+H).
Step 2: Preparation of
3-(3-(3-amino-6-(1-(1-(tert-butoxycarbonyl)piperidin-4-yl)-1H-pyrazol-4-y-
l)pyrazin-2-yl)-5-methyl-6-oxopyridazin-1(6H)-yl)-4-methylbenzoic
Acid
[0611] A solution of tert-butyl
4-(4-(5-amino-6-(1-(5-(methoxycarbonyl)-2-methylphenyl)-5-methyl-6-oxo-1,-
6-dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxy-
late (780 mg, 1.30 mmol) in 4:1 dioxane:MeOH (6.49 mL) was treated
with a solution of LiOH.H.sub.2O (136 mg, 3.25 mmol) in water (1.30
mL). The resulting mixture was stirred 4 h at ambient temperature
then concentrated under vacuum. The resulting aqueous slurry was
diluted with water (5 mL) then acidified (pH 2-3) with formic acid.
The solid formed was filtered and dried in vacuo to provide the
title compound (650 mg, 85% yield). MS (apci) m/z=587.3 (M+H).
Step 3: Preparation of tert-butyl
4-(4-(5-amino-6-(5-methyl-1-(2-methyl-5-(methylcarbamoyl)phenyl)-6-oxo-1,-
6-dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxy-
late
[0612] A cold (0.degree. C.) suspension of
3-(3-(3-amino-6-(1-(1-(tert-butoxycarbonyl)piperidin-4-yl)-1H-pyrazol-4-y-
l)pyrazin-2-yl)-5-methyl-6-oxopyridazin-1(6H)-yl)-4-methylbenzoic
acid (650 mg, 1.11 mmol) in DMF (22.2 mL) was treated sequentially
with HATU (632 mg, 1.66 mmol), 2M CH.sub.3NH.sub.2 in THF (1.11 mL,
2.22 mmol) and DIPEA (1.58 mL, 8.86 mmol). The resulting mixture
was stirred 15 min at 0.degree. C. then overnight at ambient
temperature. The reaction mixture was poured into ice water and
extracted with 5% iPrOH/DCM (3.times.50 mL). The combined organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
then concentrated under vacuum. The resulting residue was purified
by silica chromatography (5-60% acetone/DCM as the gradient eluent)
to afford the title compound (550 mg, 83% yield). MS (apci)
m/z=600.3 (M+H).
Step 4: Preparation of
3-(3-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-5-methy-
l-6-oxopyridazin-1(6H)-yl)-N,4-dimethylbenzamide Dihydrochloride
Salt
[0613] A ambient temperature solution of tert-butyl
4-(4-(5-amino-6-(5-methyl-1-(2-methyl-5-(methylcarbamoyl)phenyl)-6-oxo-1,-
6-dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxy-
late (550 mg, 0.917 mmol) in DCM (1 mL) was treated with TFA (2 mL)
then stirred 1 h at ambient temperature before concentrating to
dryness under vacuum. The resulting residue was dissolved in DCM (2
mL), treated with 4 N HCl in dioxane (2 mL), stirred for 10 min at
ambient temperature, and then concentrated to dryness under vacuum.
The resulting residue was resuspended in DCM and concentrated to
dryness under vacuum twice then dried under high vacuum to afford
the title compound (410 mg, 78% yield). MS (apci) m/z=500.2
(M+H).
Example 53
##STR00204##
[0614]
6-(2-amino-5-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3,5-dimetho-
xyphenyl)pyridazin-3 (2H)-one
[0615] A solution of
3-bromo-5-(1-methyl-1H-pyrazol-4-yl)pyrazin-2-amine (Intermediate
L20; 204.8 mg, 0.8061 mmol) in 4:1 dioxane:water (7.5 mL) was
treated with
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3 (2H)-one (Intermediate R8; 293.4 mg, 0.8192 mmol),
PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (61.27 mg, 0.07448 mmol),
K.sub.2CO.sub.3(s) (308.8 mg, 2.234 mmol). The resulting mixture
was sparged with Ar.sub.(g), then sealed and stirred for 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (50 mL), and extracted with
water (2.times.25 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The resulting crude residue was purified by C18 reverse
phase chromatography (5-95% water-ACN with 0.1% TFA as the gradient
eluent) to afford the title compound as a TFA salt. The TFA salt
was dissolved in 4:1 DCM:iPrOH (50 mL) and extracted with saturated
NaHCO.sub.3(aq) (1.times.25 mL). The organic extracts were dried
over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to afford the title compound along with impurities. The
resulting residue was subjected to further purification by silica
chromatography (40-100% DCM-Acetone as the gradient eluent) to
cleanly afford the title compound (40.1 mg, 13% yield). MS (apci)
m/z=405.1 (M+H). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
8.32-8.31 (d, 1H), 8.28-8.25 (d, 1H), 8.14-8.12 (m, 2H), 7.89 (s,
1H), 7.20-7.18 (d, 1H), 6.89 (s, 2H), 6.82-6.81 (d, 2H), 6.61-6.60
(t, 1H), 3.85 (s, 3H), 3.78 (s, 6H).
Example 54
##STR00205##
[0616]
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3-
,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0617] Step 1: Preparation of tert-butyl
4-(4-(6-amino-5-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin-3-yl)-
pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate. A solution
of tert-butyl
4-(4-(6-amino-5-bromopyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L21; 622.4 mg, 1.474 mmol) in 4:1 dioxane:water
(15 mL) was treated with
2-(3,5-dimethoxyphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)py-
ridazin-3(2H)-one (Intermediate R8; 580.7 mg, 1.621 mmol),
PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (121.2 mg, 0.1474 mmol) and
K.sub.2CO.sub.3(s) (611.0 mg, 4.421 mmol). The resulting mixture
was sparged with Ar.sub.(g), then sealed and stirred 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (250 mL) and washed with
water (2.times.50 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum. The crude residue was purified by silica chromatography
(5-95% DCM-Acetone as the gradient eluent) to afford the title
compound which was immediately carried on to step 2. MS (apci)
m/z=574.2 (M+H).
Step 2: Preparation of
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one
[0618] tert-butyl
4-(4-(6-amino-5-(1-(3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropyridazin-3-yl)-
pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate was
dissolved in 1:1 TFA:DCM (15.0 mL) then stirred 30 min at ambient
temperature. The reaction mixture was concentrated under vacuum
then purified by C18 reverse phase chromatography (5-95% ACN in
H.sub.2O with 0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (415.9 mg, 60% yield). MS (apci)
m/z=474.1 (M+H).
Example 55
##STR00206##
[0619]
6-(2-amino-5-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3--
yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0620] A solution of
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 54; 134.4 mg, 0.2838
mmol) in 1:1 DCM:MeOH (2.0 mL) was treated with formaldehyde (426.5
.mu.L, 5.677 mmol) and sodium triacetoxyborohydride (300.8 mg,
1.419 mmol) and then stirred 60 h at ambient temperature. The
reaction mixture was directly purified by silica chromatography
(1-30% DCM-MeOH w/2% NH.sub.4OH as the gradient eluent). The
chromatographic fractions containing the title compound were
combined then re-purified using C18 reverse phase chromatography
(5-95% water-ACN w/0.1% TFA as the gradient eluent) to afford the
title compound as a TFA salt. The TFA salt was dissolved in 4:1
DCM:iPrOH (50 mL) and extracted with saturated NaHCO.sub.3(aq)
(1.times.25 mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (46.9 mg, 34% yield). MS (apci)
m/z=488.2 (M+H). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
8.35-8.34 (d, 1H), 8.29 (s, 1H), 8.26 (s, 1H), 8.16-8.15 (d, 1H),
7.91 (s, 1H), 7.21-7.18 (d, 1H), 6.91 (s, 2H), 6.82-6.81 (d, 2H),
6.61-6.60 (t, 1H), 4.13-4.05 (m, 1H), 3.78 (s, 6H), 2.88-2.85 (m,
2H), 2.22 (s, 3H), 2.09-1.91 (m, 6H).
Example 56
##STR00207##
[0621]
6-(2-amino-5-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyridin-3-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0622] A solution of
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3,5-dim-
ethoxyphenyl)pyridazin-3(2H)-one (Example 54; 54.1 mg, 0.114 mmol)
in DMF (1.2 mL) was treated with 1-bromo-2-methoxyethane (21.5
.mu.L, 0.228 mmol) and K.sub.2CO.sub.3(s) (63.2 mg, 0.457 mmol).
The resulting mixture was stirred 60 h at ambient temperature. The
reaction mixture was diluted with EtOAc (25 mL) and washed with
water (2.times.10 mL) and brine (10 mL). The organic extracts were
dried over anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated
under vacuum. The crude residue was purified by silica
chromatography (1-30% DCM-MeOH w/2% NH.sub.4OH as the gradient
eluent) to afford the title compound (29.5 mg, 49% yield). MS
(apci) m/z=532.2 (M+H).
Example 57
##STR00208##
[0623]
6-(2-amino-5-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)-2-(2,6-dichlor-
o-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0624] A solution of
6-(2-amino-5-bromopyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3(2H)-one (Intermediate M2; 14.4 mg, 0.0305 mmol) in 4:1
dioxane:water (1.0 mL) was treated with
1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole
(6.98 mg, 0.0336 mmol), PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (2.51 mg,
0.00305 mmol), K.sub.2CO.sub.3(s) (12.6 mg, 0.0915 mmol). The
resulting mixture was sparged with Ar.sub.(g), then sealed and
stirred for 16 h at 100.degree. C. After cooling to ambient
temperature, the reaction mixture was diluted with EtOAc (25 mL),
and extracted with water (3.times.10 mL) and brine (10 mL). The
organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum. The resulting crude residue
was purified by silica chromatography (5-95% DCM-Acetone as the
gradient eluent) to afford the title compound (5.8 mg, 40% yield).
MS (apci) m/z=473.0 (M+H). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. 8.39-8.37 (d, 1H), 8.34-8.33 (d, 1H), 8.19-8.18 (d, 1H),
8.12 (s, 1H), 7.89 (s, 1H), 7.33-7.30 (d, 1H), 7.12 (s, 1H), 6.72
(s, 2H), 4.01 (s, 6H), 3.85 (s, 3H).
Example 58
##STR00209##
[0625]
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(2-
,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
Step 1: Preparation of Tert-Butyl
4-(4-(6-amino-5-(1-(2,6-dichloro-3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropy-
ridazin-3-yl)pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0626] A solution of
6-(2-amino-5-bromopyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3(2H)-one (Intermediate M2; 215.0 mg, 0.4554 mmol) in 4:1
dioxane:water (4.6 mL) was treated with tert-Butyl
4-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrazol-1-yl]piperidine-
-1-carboxylate (180.4 mg, 0.4782 mmol),
PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (37.46 mg, 0.04554 mmol) and
K.sub.2CO.sub.3(s) (188.8 mg, 1.366 mmol) was sparged with
Ar.sub.(g), then sealed and stirred 6 h at 90.degree. C. After
cooling to ambient temperature, the reaction mixture was diluted
with 4:1 DCM:iPrOH (50 mL) and washed with water (2.times.25 mL).
The organic extracts were dried over anhydrous Na.sub.2SO.sub.4(s),
filtered and concentrated under vacuum to afford the title compound
in sufficient purity for immediate use in step 2. MS (apci)
m/z=646.2 [(M+H)+4], 644.2 [(M+H)+2], 642.2 (M+H) with diCl
pattern.
Step 2: Preparation of
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(2,6-dic-
hloro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one
[0627] The tert-butyl
4-(4-(6-amino-5-(1-(2,6-dichloro-3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropy-
ridazin-3-yl)pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
was dissolved in 1:1 TFA:DCM (3.0 mL) then stirred 1 h at ambient
temperature. The reaction mixture was concentrated under vacuum
then purified by C18 reverse phase chromatography (2->75%
water-ACN with 0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (172.7 mg, 70% yield). MS (apci)
m/z=546.1 [(M+H)+4], 544.2 [(M+H)+2], 542.1 (M+H) with diCl
pattern.
Example 59
##STR00210##
[0628]
6-(2-amino-5-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3--
yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0629] A solution of
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(2,6-dic-
hloro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one (Example 58; 50.0 mg,
0.0922 mmol) in MeOH (1.8 mL) was treated with formaldehyde (139
.mu.L, 1.84 mmol) and stirred 15 min at ambient temperature. The
resulting reaction mixture was treated with sodium
triacetoxyborohydride (78.1 mg, 0.369 mmol) and stirred 16 h at
ambient temperature. The reaction mixture was concentrated under
vacuum then purified by C18 reverse phase chromatography (5-95%
water-ACN w/0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (24.2 mg, 47% yield). MS (apci)
m/z=558.2 [(M+H)+2], 556.2 (M+H) with diCl pattern. H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.39-8.35 (m, 2H), 8.26-8.24 (m, 1H),
8.21-8.20 (m, 1H), 7.91 (s, 1H), 7.33-7.30 (d, 1H), 7.12 (s, 1H),
6.72 (s, 2H), 4.13-4.05 (m, 1H), 4.01 (s, 6H), 2.87-2.84 (m, 2H),
2.21 (s, 3H), 2.09-1.79 (m, 6H).
Example 60
##STR00211##
[0630]
6-(2-amino-5-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyridin-3-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3
(2H)-one
[0631] A solution of
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(2,6-dic-
hloro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one (Example 58; 17 mg,
0.031 mmol) in DMF (0.31 mL) was treated with
1-bromo-2-methoxyethane (3.8 .mu.L, 0.041 mmol) and
K.sub.2CO.sub.3(s) (6.1 mg, 0.045 mmol). The resulting mixture was
stirred overnight at ambient temperature, then filtered through an
acrodisc LC 25 mm Syringe filter (with 0.45 .mu.m PVDF Membrane)
and the filtrate was purified by C18 reverse phase chromatography
(5-95% ACN in H.sub.2O with 0.1% TFA as the gradient eluent) to
afford the title compound as the TFA salt. The chromatographic
fractions containing the TFA salt of the title compound were
combined then neutralized with 1M NaOH.sub.(aq), and extracted with
DCM. The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum. The
resulting residue was purified further by silica chromatography
(10-90% CHCl.sub.3/10% MeOH/1% NH.sub.4OH in CHCl.sub.3 as the
gradient eluent) to afford the title compound (4.0 mg, 21% yield).
MS (apci) m/z=600.2 (M+H). H NMR (400 MHz, CD.sub.3OD) .delta.
8.30-8.27 (d, 1H), 8.26 (d, 1H), 8.18 (d, 1H), 8.09 (d, 1H),
8.88-8.85 (m, 2H), 7.26-7.23 (d, 1H), 7.0 (s, 1H), 4.24-4.16 (m,
1H), 4.00 (s, 6H), 3.57-3.54 (t, 3H), 3.34 (s, 3H), 3.15-3.12 (m,
2H), 2.69-2.66 (t, 2H), 2.36-2.29 (m, 2H), 2.14-2.08 (m, 4H).
Example 61
##STR00212##
[0632]
3-(3-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-6-
-oxopyridazin-1 (6H)-yl)-N-methylbenzamide
Step 1: Preparation of Tert-Butyl
4-(4-(6-amino-5-(1-(3-(methylcarbamoyl)phenyl)-6-oxo-1,6-dihydropyridazin-
-3-yl)pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
[0633] A solution of
3-(3-(2-amino-5-bromopyridin-3-yl)-6-oxopyridazin-1(6H)-yl)-N-methylbenza-
mide (Intermediate M1; 47.6 mg, 0.119 mmol) in 4:1 dioxane:water
(1.2 mL) was treated with tert-Butyl
4-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrazol-1-yl]piperidine-
-1-carboxylate (49.4 mg, 0.131 mmol),
PdCl.sub.2(dppf).CH.sub.2Cl.sub.2 (9.78 mg, 0.0119 mmol) and
K.sub.2CO.sub.3(s) (49.3 mg, 0.357 mmol). The resulting mixture was
sparged with Ar.sub.(g), then sealed and stirred 16 h at
100.degree. C. After cooling to ambient temperature, the reaction
mixture was diluted with 4:1 DCM:iPrOH (25 mL) and washed with
water (2.times.10 mL). The organic extracts were dried over
anhydrous Na.sub.2SO.sub.4(s), filtered and concentrated under
vacuum to afford the title compound which was immediately carried
on to step 2. MS (apci) m/z=571.3 (M+H).
Step 2: Preparation of
3-(3-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-6-oxopy-
ridazin-1(6H)-yl)-N-methylbenzamide
[0634] tert-Butyl
4-(4-(6-amino-5-(1-(3-(methylcarbamoyl)phenyl)-6-oxo-1,6-dihydropyridazin-
-3-yl)pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate was
dissolved in 1:1 TFA:DCM (1.2 mL) then stirred 30 min at ambient
temperature. The reaction mixture was concentrated under vacuum
then purified by C18 reverse phase chromatography (5-95% ACN in
H.sub.2O with 0.1% TFA as the gradient eluent) to afford the title
compound as a TFA salt. The TFA salt was dissolved in 4:1 DCM:iPrOH
(50 mL) and extracted with saturated NaHCO.sub.3(aq) (1.times.25
mL). The organic extracts were dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated under vacuum to
cleanly afford the title compound (18.8 mg, 34% yield). MS (apci)
m/z=471.2 (M+H).
Example 62
##STR00213##
[0635]
3-(3-(2-amino-5-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4--
yl)pyridin-3-yl)-6-oxopyridazin-1(6H)-yl)-N-methylbenzamide
2,2,2-trifluoroacetate salt
[0636] A mixture of
3-(3-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-6-oxopy-
ridazin-1(6H)-yl)-N-methylbenzamide (Example 61; 9.88 mg, 0.0210
mmol), 1-bromo-2-methoxyethane (2.17 .mu.L, 0.0231 mmol) and
K.sub.2CO.sub.3(s) (4.35 mg, 0.0315 mmol) in DMSO (0.21 mL) was
stirred overnight at ambient temperature. The reaction mixture was
diluted with DCM and washed with water and brine. The organic
extracts were dried over anhydrous Na.sub.2SO.sub.4(s), filtered
and concentrated in vacuo. The crude residue was purified by C18
reverse phase chromatography (5-95% ACN:water with 0.1% TFA) to
afford the title compound (4.8 mg, 37% yield). MS (apci) m/z=444.1
(M+H).
Example 63
##STR00214##
[0637]
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3-
-methylquinolin-7-yl)pyridazin-3 (2H)-one
bis(2,2,2-trifluoroacetate)
[0638] Step 1: Preparation of tert-butyl
4-(4-(5-amino-6-(1-(3-methylquinolin-7-yl)-6-oxo-1,6-dihydropyridazin-3-y-
l)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate: A
suspension of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 25.640 mg, 0.060570 mmol),
2-(3-methylquinolin-7-yl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-
pyridazin-3(2H)-one (Intermediate R30; 33 mg, 0.1 mmol),
Na.sub.2CO.sub.3 (90.855 .mu.L, 0.18171 mmol), and
Pd(PPh.sub.3).sub.4(7.0 mg, 0.006 mmol) in dioxane (605.70 .mu.L,
0.060570 mmol) was sparged with Ar(g), then sealed and stirred 16 h
at 90.degree. C. After cooling to ambient temperature, the reaction
mixture was partitioned between EtOAc and water. The organic
extracts were washed with brine, dried over anhydrous
Na.sub.2SO.sub.4(s), filtered and concentrated. The residue was
purified using silica gel chromatography 0-15% MeOH in EtOAc to
afford the title compound (12 mg, 0.02 mmol, 34.2% yield). MS
(apci) m/z=580.3 (M+H).
Step 2: Preparation of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3-methy-
lquinolin-7-yl)pyridazin-3(2H)-one bis(2,2,2-trifluoroacetate)
[0639] tert-butyl
4-(4-(5-amino-6-(1-(3-methylquinolin-7-yl)-6-oxo-1,6-dihydropyridazin-3-y-
l)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate (12 mg,
0.021 mmol) was dissolved in 1:1 TFA/DCM (2.0 mL) and stirred at
ambient temperature for 6 h. The reaction mixture was concentrated
under vacuum, and then treated with 2 mL MeOH. The mixture was
concentrated under vacuum to afford the title compound as the TFA
salt (9.1 mg, 0.019 mmol, 92% yield) MS (apci) m/z=480.2 (M+H).
Example 64
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-
-2-yl)-2-(3-methylquinolin-7-yl)pyridazin-3 (2H)-one
[0640] A solution
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(3-methy-
lquinolin-7-yl)pyridazin-3(2H)-one bis(2,2,2-trifluoroacetate)
(Example 63; 8 mg, 0.02 mmol) in DMSO (556 .mu.L) was treated with
K.sub.2CO.sub.3(s) (4.6 mg, 0.03 mmol). The mixture was cooled to
0.degree. C. in an ice water bath, and 1-bromo-2-methoxyethane (2
.mu.L, 0.02 mmol) was added. The resulting mixture was warmed to
ambient temperature and stirred for 50 h. The reaction was diluted
with water and extracted with DCM. The organic extracts were washed
with brine, dried over anhydrous Na.sub.2SO.sub.4(s), filtered and
concentrated under vacuum. The crude residue was purified by silica
gel chromatography (0-100% Acetone in DCM as the gradient eluent
followed by 10% MeOH in DCM) to afford the title compound (1.8 mg,
20% yield). MS (apci) m/z=538.2 (M+H).
Example 65
##STR00215##
[0641]
(S)-6-(3-amino-6-(1-((4,5,5-trimethylmorpholin-2-yl)methyl)-1H-pyra-
zol-4-yl)pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
2,2,2-trifluoroacetate
[0642] A solution of
(S)-6-(3-amino-6-(1-((5,5-dimethylmorpholin-2-yl)methyl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxyphenyl)pyridazin-3(2H)-one (29 mg,
0.0559 mmol) (Example 27; 29 mg, 0.06 mmol) in MeOH (0.5 mL) was
treated with formaldehyde (12.6 .mu.L, 0.168 mmol) and stirred for
15 min at ambient temperature. The resulting reaction mixture was
treated with sodium triacetoxyborohydride (35.6 mg, 0.168 mmol) and
stirred 30 minutes at ambient temperature. The reaction mixture was
concentrated under vacuum and then purified by C18 reverse phase
chromatography (5-95% water-ACN w/0.1% TFA as the gradient eluent)
to afford the title compound as a TFA salt (22 mg, 74% yield). MS
(apci) m/z=533.2 (M+H).
Example 66
##STR00216##
[0643]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3
(2H)-one
[0644] Step 1: A mixture of tert-butyl
4-(4-(5-amino-6-bromopyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (Intermediate L19; 0.225 g, 0.532 mmol),
2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methyl-6-(4,4,5,5-tetramethyl-1,3,-
2-dioxaborolan-2-yl)pyridazin-3 (2H)-one (Intermediate R16; 0.234
g, 0.532 mmol), Pd(PPh.sub.3).sub.4(0.0461 g, 0.0399 mmol) and 2 M
solution of Na.sub.2CO.sub.3 (0.558 mL, 1.12 mmol) in dioxane (3
mL) was stirred at 90.degree. C. for 8 hours. The mixture was then
filtered through filter paper and the filtrate was concentrated.
The residue was purified on a silica column using Hexanes:EtOAc
(10-90%) to give tert-butyl
4-(4-(5-amino-6-(1-(2,6-dichloro-3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6--
dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (0.213 g, 0.324 mmol, 60.9% yield). MS (apci) m/z=657.2
(M+H).
[0645] Step 2: A mixture of tert-butyl
4-(4-(5-amino-6-(1-(2,6-dichloro-3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6--
dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te (0.213 g, 0.324 mmol) in TFA (1.5 ml) and DCM (1.62 ml) was
stirred at room temperature for 30 min and then concentrated. The
residue was partitioned between DCM and aqueous saturated
Na.sub.2CO.sub.3. The combined organic extracts were washed with
brine, dried over Na.sub.2SO.sub.4 and concentrated to give
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,6-dic-
hloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one (0.157 g,
0.282 mmol, 86.9% yield). MS (apci) m/z=557.2 (M+H).
[0646] Step 3: 1-bromo-2-methoxyethane (0.009365 ml, 0.09965 mmol)
was added to
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-
-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one
(0.0505 g, 0.09059 mmol) and K.sub.2CO.sub.3 (0.03756 g, 0.2718
mmol) in DMF (0.9059 ml) at room temperature. The reaction mixture
was stirred for 3 days. The mixture was taken up in DCM and
extracted with water. The combined organic extracts were washed
with water and brine, dried over Na.sub.2SO.sub.4, filtered and
concentrated. The residue was purified by C18 reverse phase
chromatography (5-95% ACN:water with 0.1% TFA). The isolated
product was taken up in DCM and washed with aqueous saturated
Na.sub.2CO.sub.3 and brine, dried over Na.sub.2SO.sub.4, filtered
and concentrated to provide
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)pyrazi-
n-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one
(0.0145 g, 0.02356 mmol, 26.00% yield). MS (apci) m/z=615.2
(M+H).
Example 67
##STR00217##
[0647]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(3,5-dimethoxy-2-methylphenyl)pyridazin-3(2H)-one
[0648] Step 1: To a solution of
6-chloro-2-(3,5-dimethoxyphenyl)pyridazin-3 (2H)-one (0.505 g, 1.89
mmol) in acetonitrile (18.9 mL, 1.89 mmol) was added
1-bromopyrrolidine-2,5-dione (0.337 g, 1.89 mmol) at 0.degree. C.
The reaction mixture was warmed to ambient temperature after 10
minutes and stirred for 2.5 hours. The mixture was partitioned
between EtOAc and water. The combined organic phases were washed
with water and brine, dried over Na.sub.2SO.sub.4, filtered and
concentrated in vacuo. The residue was purified by flash
chromatography (10-90% EtOAc in hexanes) to yield
2-(2-bromo-3,5-dimethoxyphenyl)-6-chloropyridazin-3(2H)-one (0.611
g, 1.77 mmol, 93.4% yield). MS (apci) m/z=346.9 [(M+H)+2], 344.9
(M+H) with Br pattern.
[0649] Step 2: To a solution of
2-(2-bromo-3,5-dimethoxyphenyl)-6-chloropyridazin-3(2H)-one (0.308
g, 0.891 mmol) in THF (5.94 ml, 0.891 mmol) was added
methylzinc(II) chloride (0.446 mL, 0.891 mmol) and the mixture was
sparged with Ar for 15 min. Bis(tri-t-butylphosphine) Pd(0) (0.0455
g, 0.0891 mmol) was added and the reaction mixture was heated to
60.degree. C. under N2 for 3 hours. The mixture was concentrated in
vacuo and the concentrate was suspended in DCM, washed with water
and brine, dried over Na.sub.2SO.sub.4, filtered and concentrated
in vacuo. The residue was purified by flash chromatography (10-90%
EtOAc in hexanes) to yield
6-chloro-2-(3,5-dimethoxy-2-methylphenyl)pyridazin-3(2H)-one (0.109
g, 0.388 mmol, 43.6% yield). MS (apci) m/z=281.0 (M+H).
[0650] Step 3: To a solution of
6-chloro-2-(3,5-dimethoxy-2-methylphenyl)pyridazin-3(2H)-one (0.031
g, 0.11 mmol) and
4,4,4',4',5,5,5',5'-octamethyl-2,2'-bi(1,3,2-dioxaborolane) (0.042
g, 0.17 mmol) in 1,4-dioxane (1.1 mL, 0.11 mmol) was added
potassium acetate (0.033 g, 0.33 mmol) and the mixture was sparged
with Ar for 5 mins.
2-(Dicyclohexylphosphino)-2,4,6-Triisopropylbiphenyl (0.0079 g,
0.017 mmol) and Palladium(II) acetate (0.0025 g, 0.011 mmol) were
then added sequentially and the reaction mixture was sparged with
Ar. The reaction vessel was sealed and the reaction mixture was
heated to 100.degree. C. for 1 hr. The mixture was partitioned
between DCM and water. The combined organic extracts were washed
with water and brine, dried over Na.sub.2SO.sub.4, filtered and
concentrated in vacuo to provide
2-(3,5-dimethoxy-2-methylphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3(2H)-one (0.041 g, 0.11 mmol, assumed
quantitative yield).
[0651] Step 4: To a solution of
3-bromo-5-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-
-amine (Intermediate L22; 0.034 g, 0.089 mmol) and
2-(3,5-dimethoxy-2-methylphenyl)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3(2H)-one (0.040 g, 0.11 mmol) in 1,4-dioxane
(0.89 mL, 0.089 mmol) was added sodium carbonate (0.13 mL, 0.27
mmol) and the reaction mixture was sparged with Ar for 5 mins.
Tetrakis(triphenylphosphine)Pd(0) (0.0082 g, 0.0071 mmol) was added
and the reaction mixture was sparged with Ar. The mixture was
sealed and heated to 100.degree. C. for 2 hours with stirring. The
mixture was cooled to ambient temperature and purified by reverse
phase chromatography (5-95% ACN:water with 0.1% TFA) to provide the
title compound (0.020 g, 0.037 mmol, 41% yield) as a yellow powder.
MS (apci) m/z=547.3 (M+H).
Example 68
##STR00218##
[0652]
6-(3-amino-6-(1-(2-oxo-2-(piperidin-1-yl)ethyl)-1H-pyrazol-4-yl)pyr-
azin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3
(2H)-one
[0653] To a solution of
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3(2H)-one (Intermediate M3; 0.025 g, 0.053 mmol) in
1,4-dioxane (0.53 mL, 0.053 mmol) was added sodium carbonate (0.079
mL, 0.16 mmol) and
1-(piperidin-1-yl)-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H--
pyrazol-1-yl)ethanone (0.020 g, 0.063 mmol) and the reaction
mixture was sparged with Ar for 5 mins.
Tetrakis(triphenylphosphine)palladium(0) (0.0049 g, 0.0042 mmol)
was added and the reaction mixture was sparged with Ar. The
reaction was sealed, heated to 100.degree. C. and stirred for 2.5
hrs. The reaction was cooled to ambient temperature and purified by
reverse phase chromatography (5-95% ACN:water with 0.1% TFA) to
provide the title compound (0.020 g, 0.037 mmol, 41% yield) as a
yellow powder. MS (apci) m/z=589.2 [(M+H)+4], 587.2 [(M+H)+2],
585.1 (M+H) with di Cl pattern.
Example 69
##STR00219##
[0654]
6-(3-amino-6-(1-(2-(dimethylamino)ethyl)-1H-pyrazol-4-yl)pyrazin-2--
yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyridazin-3 (2H)-one
[0655] To a solution of
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3(2H)-one (Intermediate M3; 0.026 g, 0.055 mmol) in
1,4-dioxane (0.55 mL, 0.053 mmol) was added sodium carbonate (0.079
mL, 0.16 mmol) and
N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazo-
l-1-yl)ethanamine (0.019 g, 0.071 mmol) and the reaction mixture
was sparged with Ar for 5 mins.
Tetrakis(triphenylphosphine)Palladium(0) (0.0051 g, 0.0044 mmol)
was added and the reaction mixture was sparged with Ar. The
reaction was sealed, heated to 100.degree. C. and stirred for 2.5
hrs. The mixture was cooled to ambient temperature and purified by
flash chromatography (1-9% MeOH in DCM) to yield the title compound
(0.018 g, 0.034 mmol, 62% yield) as a yellow powder. MS (apci)
m/z=535.1 [(M+H)+4], 533.1 [(M+H)+2], 531.2 (M+H) with di Cl
pattern. .sup.1H NMR (DMSO) .delta. 8.71 (d, 1H), 8.48 (s, 1H),
8.32 (d, 1H), 8.00 (d, 1H), 7.30 (d, 1H), 7.10 (s, 1H), 7.03 (br s,
2H, NH.sub.2), 4.20 (t, 2H), 3.99 (s, 6H), 2.66 (t, 2H), 2.15 (s,
6H).
[0656] The following compounds shown in Table 6 were prepared
according the method used for the synthesis of Example 69 using
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)pyrid-
azin-3(2H)-one (Intermediate M3; 0.026 g, 0.055 mmol). Reaction
progression in each was followed by LCMS and reaction time was
adjusted as necessary. All compounds were purified using a method
similar to that used in either Example 68 or Example 69 utilizing
the appropriate gradient eluent. Table 6
TABLE-US-00023 TABLE 6 MS (apci) Ex. # Structure Name m/z 70
##STR00220## 6-(3-amino-6-(1-methyl- 3-(trifluoromethyl)-1H-
pyrazol-4-yl)pyrazin-2- yl)-2-(2,6-dichloro-3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 542.1 (M + H) 71 ##STR00221##
6-(3-amino-6-(1,3,5- trimethyl-1H-pyrazol-4-
yl)pyrazin-2-yl)-2-(2,6- dichloro-3,5- dimethoxyphenyl)pyridazin-
3(2H)-one 502.1 (M + H) 72 ##STR00222## 6-(3-amino-6-(1-(1-
hydroxy-2- methylpropan-2-yl)-1H- pyrazol-4-yl)pyrazin-2-
yl)-2-(2,6-dichloro-3,5- dimethoxyphenyl)pyridazin- 3(2H)-one 532.1
(M + H) 73 ##STR00223## 6-(3-amino-6-(1- isopropyl-1H-pyrazol-4-
yl)pyrazin-2-yl)-2-(2,6- dichloro-3,5- dimethoxyphenyl)pyridazin-
3(2H)-one 502.1 (M + H) 74 ##STR00224## 6-(3-amino-6-(1-
cyclopropyl-1H-pyrazol- 4-yl)pyrazin-2-yl)-2-(2,6- dichloro-3,5-
dimethoxyphenyl)pyridazin- 3(2H)-one 500.1 (M + H) 75 ##STR00225##
6-(3-amino-6-(1- (azetidin-3-yl)-1H- pyrazol-4-yl)pyrazin-2-
yl)-2-(2,6-dichloro-3,5- dimethoxyphenyl)pyridazin- 3(2H)-one 515.1
(M + H)
Example 76
##STR00226##
[0657]
6-(3-amino-6-(1-(2-(dimethylamino)ethyl)-1H-pyrazol-4-yl)pyrazin-2--
yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3
(2H)-one
[0658] To a solution of
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-me-
thylpyridazin-3(2H)-one (Intermediate M4; 0.069 g, 0.14 mmol) in
1,4-dioxane (1.42 ml, 0.142 mmol) was added sodium carbonate (0.21
mL, 0.43 mmol) and
N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazo-
l-1-yl)ethanamine (0.0488 g, 0.184 mmol) and the reaction mixture
was sparged with Ar for 5 mins.
Tetrakis(Triphenylphosphine)Palladium(0) (0.0131 g, 0.0113 mmol)
was added and the reaction mixture was sparged with Ar. The
reaction was sealed, heated to 100.degree. C. and stirred for 2
hrs. The mixture was cooled to ambient temperature and purified by
flash chromatography (1-9% MeOH in DCM) to yield the title compound
(0.0256 g, 0.0469 mmol, 33.1% yield) as a yellow powder. MS (apci)
m/z=549.1 [(M+H)+4], 547.1 [(M+H)+2], 545.2 (M+H) with di Cl
pattern. .sup.1H NMR (CDCl.sub.3) .delta. 8.48 (m, 1H), 8.27 (s,
1H), 7.96 (d, 1H), 7.94 (d, 1H), 6.71 (s, 1H), 6.20 (br s, 2H,
NH.sub.2), 4.30 (t, 2H), 3.99 (s, 6H), 2.83 (t, 2H), 2.41 (d, 3H),
2.31 (s, 6H).
Example 77
##STR00227##
[0659]
6-(3-amino-6-(1-(azetidin-3-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,-
6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one
[0660] Step 1: To a solution of
6-(3-amino-6-bromopyrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-me-
thylpyridazin-3(2H)-one (Intermediate M4; 0.120 g, 0.246 mmol) in
1,4-dioxane (2.46 ml, 0.246 mmol) was added sodium carbonate (0.370
mL, 0.739 mmol) and tert-butyl
3-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)azetid-
ine-1-carboxylate (0.103 g, 0.296 mmol) and the reaction mixture
was sparged with Ar. tetrakis(triphenylphosphine)palladium(0)
(0.0228 g, 0.0197 mmol) was added and the reaction mixture was
sparged with Ar. The reaction was sealed, heated to 100.degree. C.
and stirred for 2 hrs. The mixture was cooled to ambient
temperature and purified by flash chromatography (1-5% MeOH in DCM)
to yield the title compound (0.0256 g, 0.0469 mmol, 33.1% yield) as
a yellow powder. MS (apci) m/z=633.2 [(M+H)+4], 631.2 [(M+H)+2],
629.2 (M+H) with di Cl pattern.
[0661] Step 2: 2,2,2-Trifluoroacetic acid (1.0 ml, 0.10 mmol) was
added to a solution of tert-butyl
3-(4-(5-amino-6-(1-(2,6-dichloro-3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6--
dihydropyridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)azetidine-1-carboxylat-
e (0.065 g, 0.10 mmol) in dichloromethane (1.0 mL, 0.10 mmol) at
ambient temperature for 3.5 hrs. The mixture was partitioned
between DCM and aqueous saturated Na.sub.2CO.sub.3. The combined
organic extracts were washed with brine and concentrated to provide
the title compound (0.045 g, 0.085 mmol, 82% yield) as a yellow
solid. MS (apci) m/z=533.1 [(M+H)+4], 531.1 [(M+H)+2], 529.2 (M+H)
with di Cl pattern.
Example 78
##STR00228##
[0662]
6-(3-amino-6-(1-(1-(2-methoxyethyl)azetidin-3-yl)-1H-pyrazol-4-yl)p-
yrazin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3
(2H)-one
[0663] 1-Bromo-2-methoxyethane (0.002915 mL, 0.03138 mmol) was
added to a vial containing
6-(3-amino-6-(1-(azetidin-3-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,6-dich-
loro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one (Example 77;
0.0151 g, 0.02852 mmol) and potassium carbonate (0.005913 g,
0.04279 mmol) in DMF (0.5705 mL, 0.02852 mmol) at ambient
temperature. The mixture was stirred in sealed vial for 48 hrs. The
mixture was partitioned between DCM and water. The combined organic
extracts were washed with water and brine, dried over
Na.sub.2SO.sub.4, and concentrated in vacuo. The residue was
purified by reverse phase chromatography (5-95% ACN:water with 0.1%
TFA) to yield the title compound (0.0051 g, 0.008681 mmol, 30.44%
yield) as a yellow solid. MS (apci) m/z=591.1 [(M+H)+4], 589.1
[(M+H)+2], 587.1 (M+H) with di Cl pattern.
Example 79
##STR00229##
[0664]
6-(3-amino-6-(1-(1-(2-ethoxyethyl)azetidin-3-yl)-1H-pyrazol-4-yl)py-
razin-2-yl)-2-(2,6-dichloro-3,5-dimethoxyphenyl)-4-methylpyridazin-3
(2H)-one
[0665] 1-Bromo-2-ethoxyethane (0.003632 mL, 0.03221 mmol) was added
to a vial containing
6-(3-amino-6-(1-(azetidin-3-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,6-dich-
loro-3,5-dimethoxyphenyl)-4-methylpyridazin-3(2H)-one (Example 77;
0.0155 g, 0.02928 mmol) and potassium carbonate (0.006070 g,
0.04392 mmol) in DMF (0.9760 mL, 0.02928 mmol) at ambient
temperature. The reaction stirred in a sealed vial for 15 hrs. The
mixture was partitioned between DCM and water. The combined
organics were washed with water and brine, dried over
Na.sub.2SO.sub.4, and concentrated in vacuo. The mixture was
purified by reverse phase chromatography (5-95% ACN:water with 0.1%
TFA) to yield the title compound (0.0042 g, 0.006983 mmol, 23.85%
yield) as a yellow solid. MS (apci) m/z=605.2 [(M+H)+4], 603.2
[(M+H)+2], 601.2 (M+H) with di Cl pattern.
Example 80
##STR00230##
[0666]
3-(3-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4--
yl)pyrazin-2-yl)-5-methyl-6-oxopyridazin-1(6H)-yl)-N,4-dimethylbenzamide
[0667] To a solution of the
3-(3-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-5-methy-
l-6-oxopyridazin-1(6H)-yl)-N,4-dimethylbenzamide dihydrochloride
(50 mg, 0.087 mmol) in N,N-dimethylformamide (1747 .mu.L, 0.087
mmol) at 0.degree. C. under a nitrogen atmosphere was sequentially
added K.sub.2CO.sub.3 (49 mg, 0.35 mmol) and
1-bromo-2-methoxyethane (10 .mu.L, 0.11 mmol). The mixture was
stirred at RT overnight. The resulting mixture was diluted with 5%
IPA/DCM (50 mL) and washed with water (10 mL). The organic layer
was separated, dried (MgSO.sub.4), filtered and concentrated in
vacuo. The residue was purified by flash chromatography on silica
gel (Redi Sep 24 g) eluting with 2-20% MeOH/DCM with 2% NH.sub.4OH
to provide
3-(3-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)pyr-
azin-2-yl)-5-methyl-6-oxopyridazin-1(6H)-yl)-N,4-dimethylbenzamide
(25 mg, 51% yield) as a solid. LCMS (APCI+) m/z 558.2 (M+1),
Retention time=1.751 min.
Example 81
##STR00231##
[0668]
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3-
,5-dimethoxyphenyl)-4-methylpyridazin-3 (2H)-one
Dihydrochloride
[0669] Step 1: A glass pressure tube was charged with Intermediate
R.sup.18 [crude
2-(3,5-dimethoxyphenyl)-4-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborola-
n-2-yl)pyridazin-3(2H)-one] (660 mg, 1.77 mmol), Intermediate L21
[tert-butyl
4-(4-(6-amino-5-bromopyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxyla-
te] (749 mg, 1.77 mmol), Pd(Ph.sub.3P).sub.4 (205 mg, 0.177 mmol),
sodium carbonate 2M in water (2660 .mu.L, 5.32 mmol) and
1,4-dioxane (3546 .mu.L, 1.77 mmol). The mixture was purged with
N.sub.2 for 6 minutes. The tube was sealed with a Teflon screw cap
and heated at 90.degree. C. with vigorous stirring for 16 hours.
The mixture was cooled to RT, diluted with DCM (100 mL) and washed
with water. The organic phase was separated, dried (MgSO.sub.4),
filtered and concentrated in vacuo. The residue was purified by
flash chromatography on silica gel (Redi Sep 40 g) eluting with
1-55% acetone/hexanes (20 CV) to provide tert-butyl
4-(4-(6-amino-5-(1-(3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
(780 mg, 75% yield) as a solid. LCMS (APC1+) m/z 588.2 (M+1),
retention time=2.528 min.
[0670] Step 2: Neat TFA (3 mL) was added to the tert-butyl
4-(4-(6-amino-5-(1-(3,5-dimethoxyphenyl)-5-methyl-6-oxo-1,6-dihydropyrida-
zin-3-yl)pyridin-3-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
(600 mg, 1.02 mmol). The mixture was stirred at RT for 1 hour. The
TFA was removed in vacuo and the residue was treated with 4N HCl in
dioxane (5 mL). The mixture was stirred at RT for 15 minutes and
the solvent was removed in vacuo. The residue was evaporated from
CH.sub.3CN and dried under high vacuum to provide
6-(2-amino-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-3-yl)-2-(3,5-dim-
ethoxyphenyl)-4-methylpyridazin-3(2H)-one dihydrochloride (425 mg,
74.3% yield) as a solid. LCMS (APCI+) m/z 488.2 (M+1); Retention
time=2.261 min.
Example 82
##STR00232##
[0671]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(2,6-difluoro-3,5-dimethoxyphenyl)pyridazin-3
(2H)-one
[0672] Step 1: A mixture of Intermediate L19 (228 mg, 0.538 mmol),
Intermediate R.sup.29 (212 mg, 0.538 mmol), K.sub.2CO.sub.3 (2M,
807 .mu.L, 1.61 mmol) and Pd(Ph.sub.3P).sub.4 (31.1 mg, 0.027 mmol)
in dioxane (2.7 mL, 0.54 mmol) was sparged with nitrogen and heated
at 80.degree. C. for 3 h. The reaction mixture was partitioned
between ethyl acetate and water. The aqueous layer was extracted
with EtOAc. The combined organic layers were dried and
concentrated. The residue was purified by flash chromatography
eluting with a hexanes/EtOAc gradient (0-100%) to provide
tert-butyl
4-(4-(5-amino-6-(1-(2,6-difluoro-3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropy-
ridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate.
LCMS (APCI+) m/z 611.2 (M+1); Retention time=3.35 min.
[0673] Step 2: tert-butyl
4-(4-(5-amino-6-(1-(2,6-difluoro-3,5-dimethoxyphenyl)-6-oxo-1,6-dihydropy-
ridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
was stirred in a solution of 1:1 DCM/TFA (10 mL) for 1 h. The
mixture was concentrated at 50.degree. C. The residue was diluted
with MeOH (5 mL) and 6N HCl/iPrOH (5 mL) was added. The mixture was
concentrated to provide
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2--
(2,6-difluoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one hydrochloride
salt as a white solid. LCMS (APCI+) m/z 511.2 (M+1); Retention
time=2.35 min.
[0674] Step 3: To a suspension of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2,6-dif-
luoro-3,5-dimethoxyphenyl)pyridazin-3(2H)-one hydrochloride salt
(31.8 mg, 0.062 mmol) and K.sub.2CO.sub.3 (43.0 mg, 0.312 mmol) in
DMF (1246 .mu.L, 0.062 mmol) was added 2-bromoethyl methyl ether
(7.01 .mu.L, 0.075 mmol) and the resulting mixture was stirred at
RT for 1 d. The crude reaction mixture was loaded onto a silica
column equilibrated with hexanes and eluted with
hexanes>DCM>20% MeOH/DCM to provide the title compound (18
mg, 0.031 mmol, 50.8%). LCMS (APCI+) m/z=569.2; retention time 2.40
min.
Example 83
##STR00233##
[0675]
6-(3-amino-6-(1-(1-(2-methoxyethyl)piperidin-4-yl)-1H-pyrazol-4-yl)-
pyrazin-2-yl)-2-(2-chloro-6-fluoro-3-methoxyphenyl)pyridazin-3
(2H)-one
[0676] Step 1: A mixture of Intermediate L19 (262 mg, 0.620 mmol),
Intermediate R.sup.28 (236 mg, 0.620 mmol), K.sub.2CO.sub.3 (2 M,
930 .mu.L, 1.86 mmol) and Pd(Ph.sub.3P).sub.4 (35.8 mg, 0.031 mmol)
in dioxane (3.1 mL, 0.62 mmol) was sparged with nitrogen and heated
at 80.degree. C. for 3 h. The reaction was partitioned between
ethyl acetate and water. The combined organic layers were dried
over Na.sub.2SO.sub.4, filtered and concentrated. The residue was
purified by flash chromatography eluting with a hexane/Ethyl
acetate gradient of 0-100% to provide tert-butyl
4-(4-(5-amino-6-(1-(2-chloro-6-fluoro-3-methoxyphenyl)-6-oxo-1,6-dihydrop-
yridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate.
LCMS (APCI+) m/z=597.2 (100%), 599.2(40%); retention time 3.38
min.
[0677] Step 2: tert-butyl
4-(4-(5-amino-6-(1-(2-chloro-6-fluoro-3-methoxyphenyl)-6-oxo-1,6-dihydrop-
yridazin-3-yl)pyrazin-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate
was treated with 1:1 DCM/TFA (20 mL) for 1 h. The solution was
concentrated and 5 mL of 6N HCl/iPrOH and 5 mL of MeOH was added.
The mixture was stirred for 1 h to form a white slurry and the
suspension was concentrated. LCMS (APCI+) m/z=497.0 (100%), 499.0
(40%); retention time 2.33 min.
[0678] Step 3: To a suspension of
6-(3-amino-6-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyrazin-2-yl)-2-(2-chlor-
o-6-fluoro-3-methoxyphenyl)pyridazin-3(2H)-one (25.3 mg, 0.051
mmol) and K.sub.2CO.sub.3 (35.2 mg, 0.255 mmol) in DMF (1.0 ml,
0.051 mmol) was added 2-bromoethyl methyl ether (5.74 al, 0.06
mmol) and the resulting mixture was stirred at RT for 1 d. The
crude reaction mixture was loaded onto a silica column equilibrated
with hexanes and eluted with hexanes>DCM>20% MeOH/DCM to
provide the title compound (12 mg, 0.021 mmol, 42%). LCMS (APCI+)
m/z=555.2; retention time 2.388.
Biological Activity
Example A
Enzyme Assay
[0679] FGFR1, 2 and 3 kinase activity was measured by the
Invitrogen LanthaScreen.TM. Assay technology which directly
measures the amount of substrate phosphorylation by TR-FRET using a
fluorescein-labeled peptide and Europium-labeled antibody.
[0680] To measure FGFR1 kinase activity, 200 pM His-tagged
recombinant human FGFR1 catalytic domain (amino acids 308-731)
(Life Technologies Cat. No. PR4660A) was incubated with 100 nM
Alexa Fluor.RTM. 647-Poly-GT Peptide Substrate (Life Technologies
Cat. No. PV5836) and ATP in the presence of Mg.sup.++, along with
test compound in a buffer consisting of 250 mM HEPES, 25 mM
MgCl.sub.2, 0.05% TritonX-100, pH 7.5, and 2% DMSO. Compounds were
typically prepared in a threefold serial dilution in DMSO and added
to the assay to give the appropriate final concentration. After 20
minutes incubation at 22.degree. C., an equal volume of 2 nM
LanthaScreen.RTM. Eu-PY20 Antibody (Life Technologies Cat. No.
PV5691) and EDTA were added to quench the kinase reaction and start
the detection reaction. After an additional 60 minute incubation at
22.degree. C., the reaction was measured using a PerkinElmer
EnVision multimode plate reader via TR-FRET dual wavelength
detection, and the percent of control (POC) calculated using a
ratiometric emission factor. 100 POC is determined using no test
compound and 0 POC is determined using no enzyme. The POC values
were fit to a 4 parameter logistic curve and the IC.sub.50 value is
point where the curve crosses 50 POC.
[0681] To measure FGFR2 kinase activity: 200 pM His-tagged
recombinant human FGFR2 cytoplasmic domain (amino acids 403-822),
(Life Technologies Cat. No. PR5332A); 20 minutes incubation at
22.degree. C., 60 minute detection incubation at 22.degree. C.
[0682] To measure FGFR3 kinase activity: 750 pM N-terminal GST-HIS6
fusion protein with a 3C cleavage site recombinant human FGFR3
(amino acids R397-T806) (ProQinase Cat. No. 1068-0000-1); 10
minutes incubation at 22.degree. C., 60 minute detection incubation
at 22.degree. C.
[0683] The averaged IC.sub.50 values for the compounds tested in
this assay are provided in Table F.
TABLE-US-00024 TABLE F IC.sub.50's of compounds tested in the assay
of Example A FGFR1 Enz FRET FGFR2 Enz FRET FGFR3 Enz FRET IC50 (nM)
IC50 (nM) IC50 (nM) Ex. # [AVERAGE] [AVERAGE] [AVERAGE] 1 18.1 3.4
4.3 2 259.6 60.4 69.1 3 4749.7 1963.4 4121.7 4 15.4 2.8 2.4 5 17.0
3.2 2.9 6 22.2 6.4 11.2 7 14.5 6.5 10.7 8 N/A 120.0 176.7 9 N/A
1055.6 4251.3 10 N/A 214.7 293.0 11 143.2 23.7 22.6 12 838.2 150.6
191.5 13 316.3 64.3 15.1 14 6.3 1.6 1.7 15 17.4 3.7 7.0 16 22.8 5.6
10.6 17 79.2 10.2 3.8 18 47.7 8.5 8.7 19 15.1 2.8 2.2 20 399.9 39.8
61.5 21 39.1 7.5 4.9 22 192.5 21.1 6.7 23 3.8 5.5 2.8 24 15.0 1.8
4.0 25 51.1 6.6 11.8 26 8.6 2.2 4.3 27 19.0 1.9 8.2 28 17.0 2.3 4.7
29 22.0 3.7 7.1 30 14.1 2.9 6.5 31 7.7 1.4 3.9 32 9.2 1.3 1.8 33
11.6 1.8 4.5 34 38.4 3.7 13.4 35 15.4 3.2 4.8 36 62.8 6.2 21.3 37
56.0 19.2 23.5 38 91.5 16.8 21.1 39 141.8 9.4 28.6 40 49.7 2.9 10.2
41 3.0 0.8 3.2 42 6.3 1.4 8.6 43 7.0 1.6 1.7 44 7.1 1.4 6.2 45
1561.6 825.7 1086.6 46 3101.2 1231.5 3604.1 47 5.6 1.6 1.5 48 3.1
1.0 1.4 49 6.0 1.3 2.1 50 3.5 0.9 1.8 51 4.0 0.6 1.0 52 26.8 5.0
42.5 53 193.4 72.3 35.6 54 29.6 10.9 10.1 55 31.4 6.3 19.9 56 43.3
7.5 5.7 57 13.8 5.8 6.2 58 4.4 2.4 0.7 59 5.0 6.6 10.6 60 11.6 1.5
10.3 61 98.2 24.1 13.7 62 231.3 31.2 166.2 63 14.0 16.0 53.5 64
61.4 42.1 213.7 65 8.9 2.9 35.0 66 5.6 7.8 3.9 67 9.1 6.5 13.3 68
9.8 11.1 6.4 69 7.2 9.6 9.6 70 30.9 54.4 30.8 71 26.5 38.8 45.1 72
7.3 9.3 5.2 73 7.2 10.6 6.0 74 5.8 9.5 6.1 75 8.0 11.3 10.2 76 10.6
3.85 7.1 77 16.1 6.2 12.4 78 9.8 3.7 6.9 79 17.3 6.3 10.7 80 57.0
16.3 50.6 81 3.6 1.1 11.4 82 2.4 3.1 2.2 83 6.9 16.6 26.5 N/A = Not
available
EQUIVALENTS
[0684] Those skilled in the art will recognize, or be able to
ascertain using no more than routine experimentation, many
equivalents to the specific embodiments of the invention described
herein. Such equivalents are intended to be encompassed by the
following claims.
INCORPORATION BY REFERENCE
[0685] The entire contents of all patents, published patent
applications, websites, and other references cited herein are
hereby expressly incorporated herein in their entireties by
reference.
Sequence CWU 1
1
81822PRTHomo sapiens 1Met Trp Ser Trp Lys Cys Leu Leu Phe Trp Ala
Val Leu Val Thr Ala1 5 10 15Thr Leu Cys Thr Ala Arg Pro Ser Pro Thr
Leu Pro Glu Gln Ala Gln 20 25 30Pro Trp Gly Ala Pro Val Glu Val Glu
Ser Phe Leu Val His Pro Gly 35 40 45Asp Leu Leu Gln Leu Arg Cys Arg
Leu Arg Asp Asp Val Gln Ser Ile 50 55 60Asn Trp Leu Arg Asp Gly Val
Gln Leu Ala Glu Ser Asn Arg Thr Arg65 70 75 80Ile Thr Gly Glu Glu
Val Glu Val Gln Asp Ser Val Pro Ala Asp Ser 85 90 95Gly Leu Tyr Ala
Cys Val Thr Ser Ser Pro Ser Gly Ser Asp Thr Thr 100 105 110Tyr Phe
Ser Val Asn Val Ser Asp Ala Leu Pro Ser Ser Glu Asp Asp 115 120
125Asp Asp Asp Asp Asp Ser Ser Ser Glu Glu Lys Glu Thr Asp Asn Thr
130 135 140Lys Pro Asn Arg Met Pro Val Ala Pro Tyr Trp Thr Ser Pro
Glu Lys145 150 155 160Met Glu Lys Lys Leu His Ala Val Pro Ala Ala
Lys Thr Val Lys Phe 165 170 175Lys Cys Pro Ser Ser Gly Thr Pro Asn
Pro Thr Leu Arg Trp Leu Lys 180 185 190Asn Gly Lys Glu Phe Lys Pro
Asp His Arg Ile Gly Gly Tyr Lys Val 195 200 205Arg Tyr Ala Thr Trp
Ser Ile Ile Met Asp Ser Val Val Pro Ser Asp 210 215 220Lys Gly Asn
Tyr Thr Cys Ile Val Glu Asn Glu Tyr Gly Ser Ile Asn225 230 235
240His Thr Tyr Gln Leu Asp Val Val Glu Arg Ser Pro His Arg Pro Ile
245 250 255Leu Gln Ala Gly Leu Pro Ala Asn Lys Thr Val Ala Leu Gly
Ser Asn 260 265 270Val Glu Phe Met Cys Lys Val Tyr Ser Asp Pro Gln
Pro His Ile Gln 275 280 285Trp Leu Lys His Ile Glu Val Asn Gly Ser
Lys Ile Gly Pro Asp Asn 290 295 300Leu Pro Tyr Val Gln Ile Leu Lys
Thr Ala Gly Val Asn Thr Thr Asp305 310 315 320Lys Glu Met Glu Val
Leu His Leu Arg Asn Val Ser Phe Glu Asp Ala 325 330 335Gly Glu Tyr
Thr Cys Leu Ala Gly Asn Ser Ile Gly Leu Ser His His 340 345 350Ser
Ala Trp Leu Thr Val Leu Glu Ala Leu Glu Glu Arg Pro Ala Val 355 360
365Met Thr Ser Pro Leu Tyr Leu Glu Ile Ile Ile Tyr Cys Thr Gly Ala
370 375 380Phe Leu Ile Ser Cys Met Val Gly Ser Val Ile Val Tyr Lys
Met Lys385 390 395 400Ser Gly Thr Lys Lys Ser Asp Phe His Ser Gln
Met Ala Val His Lys 405 410 415Leu Ala Lys Ser Ile Pro Leu Arg Arg
Gln Val Thr Val Ser Ala Asp 420 425 430Ser Ser Ala Ser Met Asn Ser
Gly Val Leu Leu Val Arg Pro Ser Arg 435 440 445Leu Ser Ser Ser Gly
Thr Pro Met Leu Ala Gly Val Ser Glu Tyr Glu 450 455 460Leu Pro Glu
Asp Pro Arg Trp Glu Leu Pro Arg Asp Arg Leu Val Leu465 470 475
480Gly Lys Pro Leu Gly Glu Gly Cys Phe Gly Gln Val Val Leu Ala Glu
485 490 495Ala Ile Gly Leu Asp Lys Asp Lys Pro Asn Arg Val Thr Lys
Val Ala 500 505 510Val Lys Met Leu Lys Ser Asp Ala Thr Glu Lys Asp
Leu Ser Asp Leu 515 520 525Ile Ser Glu Met Glu Met Met Lys Met Ile
Gly Lys His Lys Asn Ile 530 535 540Ile Asn Leu Leu Gly Ala Cys Thr
Gln Asp Gly Pro Leu Tyr Val Ile545 550 555 560Val Glu Tyr Ala Ser
Lys Gly Asn Leu Arg Glu Tyr Leu Gln Ala Arg 565 570 575Arg Pro Pro
Gly Leu Glu Tyr Cys Tyr Asn Pro Ser His Asn Pro Glu 580 585 590Glu
Gln Leu Ser Ser Lys Asp Leu Val Ser Cys Ala Tyr Gln Val Ala 595 600
605Arg Gly Met Glu Tyr Leu Ala Ser Lys Lys Cys Ile His Arg Asp Leu
610 615 620Ala Ala Arg Asn Val Leu Val Thr Glu Asp Asn Val Met Lys
Ile Ala625 630 635 640Asp Phe Gly Leu Ala Arg Asp Ile His His Ile
Asp Tyr Tyr Lys Lys 645 650 655Thr Thr Asn Gly Arg Leu Pro Val Lys
Trp Met Ala Pro Glu Ala Leu 660 665 670Phe Asp Arg Ile Tyr Thr His
Gln Ser Asp Val Trp Ser Phe Gly Val 675 680 685Leu Leu Trp Glu Ile
Phe Thr Leu Gly Gly Ser Pro Tyr Pro Gly Val 690 695 700Pro Val Glu
Glu Leu Phe Lys Leu Leu Lys Glu Gly His Arg Met Asp705 710 715
720Lys Pro Ser Asn Cys Thr Asn Glu Leu Tyr Met Met Met Arg Asp Cys
725 730 735Trp His Ala Val Pro Ser Gln Arg Pro Thr Phe Lys Gln Leu
Val Glu 740 745 750Asp Leu Asp Arg Ile Val Ala Leu Thr Ser Asn Gln
Glu Tyr Leu Asp 755 760 765Leu Ser Met Pro Leu Asp Gln Tyr Ser Pro
Ser Phe Pro Asp Thr Arg 770 775 780Ser Ser Thr Cys Ser Ser Gly Glu
Asp Ser Val Phe Ser His Glu Pro785 790 795 800Leu Pro Glu Glu Pro
Cys Leu Pro Arg His Pro Ala Gln Leu Ala Asn 805 810 815Gly Gly Leu
Lys Arg Arg 8202820PRTHomo sapiens 2Met Trp Ser Trp Lys Cys Leu Leu
Phe Trp Ala Val Leu Val Thr Ala1 5 10 15Thr Leu Cys Thr Ala Arg Pro
Ser Pro Thr Leu Pro Glu Gln Ala Gln 20 25 30Pro Trp Gly Ala Pro Val
Glu Val Glu Ser Phe Leu Val His Pro Gly 35 40 45Asp Leu Leu Gln Leu
Arg Cys Arg Leu Arg Asp Asp Val Gln Ser Ile 50 55 60Asn Trp Leu Arg
Asp Gly Val Gln Leu Ala Glu Ser Asn Arg Thr Arg65 70 75 80Ile Thr
Gly Glu Glu Val Glu Val Gln Asp Ser Val Pro Ala Asp Ser 85 90 95Gly
Leu Tyr Ala Cys Val Thr Ser Ser Pro Ser Gly Ser Asp Thr Thr 100 105
110Tyr Phe Ser Val Asn Val Ser Asp Ala Leu Pro Ser Ser Glu Asp Asp
115 120 125Asp Asp Asp Asp Asp Ser Ser Ser Glu Glu Lys Glu Thr Asp
Asn Thr 130 135 140Lys Pro Asn Pro Val Ala Pro Tyr Trp Thr Ser Pro
Glu Lys Met Glu145 150 155 160Lys Lys Leu His Ala Val Pro Ala Ala
Lys Thr Val Lys Phe Lys Cys 165 170 175Pro Ser Ser Gly Thr Pro Asn
Pro Thr Leu Arg Trp Leu Lys Asn Gly 180 185 190Lys Glu Phe Lys Pro
Asp His Arg Ile Gly Gly Tyr Lys Val Arg Tyr 195 200 205Ala Thr Trp
Ser Ile Ile Met Asp Ser Val Val Pro Ser Asp Lys Gly 210 215 220Asn
Tyr Thr Cys Ile Val Glu Asn Glu Tyr Gly Ser Ile Asn His Thr225 230
235 240Tyr Gln Leu Asp Val Val Glu Arg Ser Pro His Arg Pro Ile Leu
Gln 245 250 255Ala Gly Leu Pro Ala Asn Lys Thr Val Ala Leu Gly Ser
Asn Val Glu 260 265 270Phe Met Cys Lys Val Tyr Ser Asp Pro Gln Pro
His Ile Gln Trp Leu 275 280 285Lys His Ile Glu Val Asn Gly Ser Lys
Ile Gly Pro Asp Asn Leu Pro 290 295 300Tyr Val Gln Ile Leu Lys Thr
Ala Gly Val Asn Thr Thr Asp Lys Glu305 310 315 320Met Glu Val Leu
His Leu Arg Asn Val Ser Phe Glu Asp Ala Gly Glu 325 330 335Tyr Thr
Cys Leu Ala Gly Asn Ser Ile Gly Leu Ser His His Ser Ala 340 345
350Trp Leu Thr Val Leu Glu Ala Leu Glu Glu Arg Pro Ala Val Met Thr
355 360 365Ser Pro Leu Tyr Leu Glu Ile Ile Ile Tyr Cys Thr Gly Ala
Phe Leu 370 375 380Ile Ser Cys Met Val Gly Ser Val Ile Val Tyr Lys
Met Lys Ser Gly385 390 395 400Thr Lys Lys Ser Asp Phe His Ser Gln
Met Ala Val His Lys Leu Ala 405 410 415Lys Ser Ile Pro Leu Arg Arg
Gln Val Thr Val Ser Ala Asp Ser Ser 420 425 430Ala Ser Met Asn Ser
Gly Val Leu Leu Val Arg Pro Ser Arg Leu Ser 435 440 445Ser Ser Gly
Thr Pro Met Leu Ala Gly Val Ser Glu Tyr Glu Leu Pro 450 455 460Glu
Asp Pro Arg Trp Glu Leu Pro Arg Asp Arg Leu Val Leu Gly Lys465 470
475 480Pro Leu Gly Glu Gly Cys Phe Gly Gln Val Val Leu Ala Glu Ala
Ile 485 490 495Gly Leu Asp Lys Asp Lys Pro Asn Arg Val Thr Lys Val
Ala Val Lys 500 505 510Met Leu Lys Ser Asp Ala Thr Glu Lys Asp Leu
Ser Asp Leu Ile Ser 515 520 525Glu Met Glu Met Met Lys Met Ile Gly
Lys His Lys Asn Ile Ile Asn 530 535 540Leu Leu Gly Ala Cys Thr Gln
Asp Gly Pro Leu Tyr Val Ile Val Glu545 550 555 560Tyr Ala Ser Lys
Gly Asn Leu Arg Glu Tyr Leu Gln Ala Arg Arg Pro 565 570 575Pro Gly
Leu Glu Tyr Cys Tyr Asn Pro Ser His Asn Pro Glu Glu Gln 580 585
590Leu Ser Ser Lys Asp Leu Val Ser Cys Ala Tyr Gln Val Ala Arg Gly
595 600 605Met Glu Tyr Leu Ala Ser Lys Lys Cys Ile His Arg Asp Leu
Ala Ala 610 615 620Arg Asn Val Leu Val Thr Glu Asp Asn Val Met Lys
Ile Ala Asp Phe625 630 635 640Gly Leu Ala Arg Asp Ile His His Ile
Asp Tyr Tyr Lys Lys Thr Thr 645 650 655Asn Gly Arg Leu Pro Val Lys
Trp Met Ala Pro Glu Ala Leu Phe Asp 660 665 670Arg Ile Tyr Thr His
Gln Ser Asp Val Trp Ser Phe Gly Val Leu Leu 675 680 685Trp Glu Ile
Phe Thr Leu Gly Gly Ser Pro Tyr Pro Gly Val Pro Val 690 695 700Glu
Glu Leu Phe Lys Leu Leu Lys Glu Gly His Arg Met Asp Lys Pro705 710
715 720Ser Asn Cys Thr Asn Glu Leu Tyr Met Met Met Arg Asp Cys Trp
His 725 730 735Ala Val Pro Ser Gln Arg Pro Thr Phe Lys Gln Leu Val
Glu Asp Leu 740 745 750Asp Arg Ile Val Ala Leu Thr Ser Asn Gln Glu
Tyr Leu Asp Leu Ser 755 760 765Met Pro Leu Asp Gln Tyr Ser Pro Ser
Phe Pro Asp Thr Arg Ser Ser 770 775 780Thr Cys Ser Ser Gly Glu Asp
Ser Val Phe Ser His Glu Pro Leu Pro785 790 795 800Glu Glu Pro Cys
Leu Pro Arg His Pro Ala Gln Leu Ala Asn Gly Gly 805 810 815Leu Lys
Arg Arg 8203821PRTHomo sapiens 3Met Val Ser Trp Gly Arg Phe Ile Cys
Leu Val Val Val Thr Met Ala1 5 10 15Thr Leu Ser Leu Ala Arg Pro Ser
Phe Ser Leu Val Glu Asp Thr Thr 20 25 30Leu Glu Pro Glu Glu Pro Pro
Thr Lys Tyr Gln Ile Ser Gln Pro Glu 35 40 45Val Tyr Val Ala Ala Pro
Gly Glu Ser Leu Glu Val Arg Cys Leu Leu 50 55 60Lys Asp Ala Ala Val
Ile Ser Trp Thr Lys Asp Gly Val His Leu Gly65 70 75 80Pro Asn Asn
Arg Thr Val Leu Ile Gly Glu Tyr Leu Gln Ile Lys Gly 85 90 95Ala Thr
Pro Arg Asp Ser Gly Leu Tyr Ala Cys Thr Ala Ser Arg Thr 100 105
110Val Asp Ser Glu Thr Trp Tyr Phe Met Val Asn Val Thr Asp Ala Ile
115 120 125Ser Ser Gly Asp Asp Glu Asp Asp Thr Asp Gly Ala Glu Asp
Phe Val 130 135 140Ser Glu Asn Ser Asn Asn Lys Arg Ala Pro Tyr Trp
Thr Asn Thr Glu145 150 155 160Lys Met Glu Lys Arg Leu His Ala Val
Pro Ala Ala Asn Thr Val Lys 165 170 175Phe Arg Cys Pro Ala Gly Gly
Asn Pro Met Pro Thr Met Arg Trp Leu 180 185 190Lys Asn Gly Lys Glu
Phe Lys Gln Glu His Arg Ile Gly Gly Tyr Lys 195 200 205Val Arg Asn
Gln His Trp Ser Leu Ile Met Glu Ser Val Val Pro Ser 210 215 220Asp
Lys Gly Asn Tyr Thr Cys Val Val Glu Asn Glu Tyr Gly Ser Ile225 230
235 240Asn His Thr Tyr His Leu Asp Val Val Glu Arg Ser Pro His Arg
Pro 245 250 255Ile Leu Gln Ala Gly Leu Pro Ala Asn Ala Ser Thr Val
Val Gly Gly 260 265 270Asp Val Glu Phe Val Cys Lys Val Tyr Ser Asp
Ala Gln Pro His Ile 275 280 285Gln Trp Ile Lys His Val Glu Lys Asn
Gly Ser Lys Tyr Gly Pro Asp 290 295 300Gly Leu Pro Tyr Leu Lys Val
Leu Lys Ala Ala Gly Val Asn Thr Thr305 310 315 320Asp Lys Glu Ile
Glu Val Leu Tyr Ile Arg Asn Val Thr Phe Glu Asp 325 330 335Ala Gly
Glu Tyr Thr Cys Leu Ala Gly Asn Ser Ile Gly Ile Ser Phe 340 345
350His Ser Ala Trp Leu Thr Val Leu Pro Ala Pro Gly Arg Glu Lys Glu
355 360 365Ile Thr Ala Ser Pro Asp Tyr Leu Glu Ile Ala Ile Tyr Cys
Ile Gly 370 375 380Val Phe Leu Ile Ala Cys Met Val Val Thr Val Ile
Leu Cys Arg Met385 390 395 400Lys Asn Thr Thr Lys Lys Pro Asp Phe
Ser Ser Gln Pro Ala Val His 405 410 415Lys Leu Thr Lys Arg Ile Pro
Leu Arg Arg Gln Val Thr Val Ser Ala 420 425 430Glu Ser Ser Ser Ser
Met Asn Ser Asn Thr Pro Leu Val Arg Ile Thr 435 440 445Thr Arg Leu
Ser Ser Thr Ala Asp Thr Pro Met Leu Ala Gly Val Ser 450 455 460Glu
Tyr Glu Leu Pro Glu Asp Pro Lys Trp Glu Phe Pro Arg Asp Lys465 470
475 480Leu Thr Leu Gly Lys Pro Leu Gly Glu Gly Cys Phe Gly Gln Val
Val 485 490 495Met Ala Glu Ala Val Gly Ile Asp Lys Asp Lys Pro Lys
Glu Ala Val 500 505 510Thr Val Ala Val Lys Met Leu Lys Asp Asp Ala
Thr Glu Lys Asp Leu 515 520 525Ser Asp Leu Val Ser Glu Met Glu Met
Met Lys Met Ile Gly Lys His 530 535 540Lys Asn Ile Ile Asn Leu Leu
Gly Ala Cys Thr Gln Asp Gly Pro Leu545 550 555 560Tyr Val Ile Val
Glu Tyr Ala Ser Lys Gly Asn Leu Arg Glu Tyr Leu 565 570 575Arg Ala
Arg Arg Pro Pro Gly Met Glu Tyr Ser Tyr Asp Ile Asn Arg 580 585
590Val Pro Glu Glu Gln Met Thr Phe Lys Asp Leu Val Ser Cys Thr Tyr
595 600 605Gln Leu Ala Arg Gly Met Glu Tyr Leu Ala Ser Gln Lys Cys
Ile His 610 615 620Arg Asp Leu Ala Ala Arg Asn Val Leu Val Thr Glu
Asn Asn Val Met625 630 635 640Lys Ile Ala Asp Phe Gly Leu Ala Arg
Asp Ile Asn Asn Ile Asp Tyr 645 650 655Tyr Lys Lys Thr Thr Asn Gly
Arg Leu Pro Val Lys Trp Met Ala Pro 660 665 670Glu Ala Leu Phe Asp
Arg Val Tyr Thr His Gln Ser Asp Val Trp Ser 675 680 685Phe Gly Val
Leu Met Trp Glu Ile Phe Thr Leu Gly Gly Ser Pro Tyr 690 695 700Pro
Gly Ile Pro Val Glu Glu Leu Phe Lys Leu Leu Lys Glu Gly His705 710
715 720Arg Met Asp Lys Pro Ala Asn Cys Thr Asn Glu Leu Tyr Met Met
Met 725 730 735Arg Asp Cys Trp His Ala Val Pro Ser Gln Arg Pro Thr
Phe Lys Gln 740 745 750Leu Val Glu Asp Leu Asp Arg Ile Leu Thr Leu
Thr Thr Asn Glu Glu 755 760 765Tyr Leu Asp Leu Ser Gln Pro Leu Glu
Gln Tyr Ser Pro Ser Tyr Pro 770 775 780Asp Thr Arg Ser Ser Cys Ser
Ser Gly Asp Asp Ser Val Phe Ser Pro785 790 795 800Asp Pro Met Pro
Tyr Glu Pro Cys Leu Pro Gln Tyr Pro His Ile Asn 805 810 815Gly Ser
Val Lys Thr 8204822PRTHomo sapiens 4Met Val
Ser Trp Gly Arg Phe Ile Cys Leu Val Val Val Thr Met Ala1 5 10 15Thr
Leu Ser Leu Ala Arg Pro Ser Phe Ser Leu Val Glu Asp Thr Thr 20 25
30Leu Glu Pro Glu Glu Pro Pro Thr Lys Tyr Gln Ile Ser Gln Pro Glu
35 40 45Val Tyr Val Ala Ala Pro Gly Glu Ser Leu Glu Val Arg Cys Leu
Leu 50 55 60Lys Asp Ala Ala Val Ile Ser Trp Thr Lys Asp Gly Val His
Leu Gly65 70 75 80Pro Asn Asn Arg Thr Val Leu Ile Gly Glu Tyr Leu
Gln Ile Lys Gly 85 90 95Ala Thr Pro Arg Asp Ser Gly Leu Tyr Ala Cys
Thr Ala Ser Arg Thr 100 105 110Val Asp Ser Glu Thr Trp Tyr Phe Met
Val Asn Val Thr Asp Ala Ile 115 120 125Ser Ser Gly Asp Asp Glu Asp
Asp Thr Asp Gly Ala Glu Asp Phe Val 130 135 140Ser Glu Asn Ser Asn
Asn Lys Arg Ala Pro Tyr Trp Thr Asn Thr Glu145 150 155 160Lys Met
Glu Lys Arg Leu His Ala Val Pro Ala Ala Asn Thr Val Lys 165 170
175Phe Arg Cys Pro Ala Gly Gly Asn Pro Met Pro Thr Met Arg Trp Leu
180 185 190Lys Asn Gly Lys Glu Phe Lys Gln Glu His Arg Ile Gly Gly
Tyr Lys 195 200 205Val Arg Asn Gln His Trp Ser Leu Ile Met Glu Ser
Val Val Pro Ser 210 215 220Asp Lys Gly Asn Tyr Thr Cys Val Val Glu
Asn Glu Tyr Gly Ser Ile225 230 235 240Asn His Thr Tyr His Leu Asp
Val Val Glu Arg Ser Pro His Arg Pro 245 250 255Ile Leu Gln Ala Gly
Leu Pro Ala Asn Ala Ser Thr Val Val Gly Gly 260 265 270Asp Val Glu
Phe Val Cys Lys Val Tyr Ser Asp Ala Gln Pro His Ile 275 280 285Gln
Trp Ile Lys His Val Glu Lys Asn Gly Ser Lys Tyr Gly Pro Asp 290 295
300Gly Leu Pro Tyr Leu Lys Val Leu Lys His Ser Gly Ile Asn Ser
Ser305 310 315 320Asn Ala Glu Val Leu Ala Leu Phe Asn Val Thr Glu
Ala Asp Ala Gly 325 330 335Glu Tyr Ile Cys Lys Val Ser Asn Tyr Ile
Gly Gln Ala Asn Gln Ser 340 345 350Ala Trp Leu Thr Val Leu Pro Lys
Gln Gln Ala Pro Gly Arg Glu Lys 355 360 365Glu Ile Thr Ala Ser Pro
Asp Tyr Leu Glu Ile Ala Ile Tyr Cys Ile 370 375 380Gly Val Phe Leu
Ile Ala Cys Met Val Val Thr Val Ile Leu Cys Arg385 390 395 400Met
Lys Asn Thr Thr Lys Lys Pro Asp Phe Ser Ser Gln Pro Ala Val 405 410
415His Lys Leu Thr Lys Arg Ile Pro Leu Arg Arg Gln Val Thr Val Ser
420 425 430Ala Glu Ser Ser Ser Ser Met Asn Ser Asn Thr Pro Leu Val
Arg Ile 435 440 445Thr Thr Arg Leu Ser Ser Thr Ala Asp Thr Pro Met
Leu Ala Gly Val 450 455 460Ser Glu Tyr Glu Leu Pro Glu Asp Pro Lys
Trp Glu Phe Pro Arg Asp465 470 475 480Lys Leu Thr Leu Gly Lys Pro
Leu Gly Glu Gly Cys Phe Gly Gln Val 485 490 495Val Met Ala Glu Ala
Val Gly Ile Asp Lys Asp Lys Pro Lys Glu Ala 500 505 510Val Thr Val
Ala Val Lys Met Leu Lys Asp Asp Ala Thr Glu Lys Asp 515 520 525Leu
Ser Asp Leu Val Ser Glu Met Glu Met Met Lys Met Ile Gly Lys 530 535
540His Lys Asn Ile Ile Asn Leu Leu Gly Ala Cys Thr Gln Asp Gly
Pro545 550 555 560Leu Tyr Val Ile Val Glu Tyr Ala Ser Lys Gly Asn
Leu Arg Glu Tyr 565 570 575Leu Arg Ala Arg Arg Pro Pro Gly Met Glu
Tyr Ser Tyr Asp Ile Asn 580 585 590Arg Val Pro Glu Glu Gln Met Thr
Phe Lys Asp Leu Val Ser Cys Thr 595 600 605Tyr Gln Leu Ala Arg Gly
Met Glu Tyr Leu Ala Ser Gln Lys Cys Ile 610 615 620His Arg Asp Leu
Ala Ala Arg Asn Val Leu Val Thr Glu Asn Asn Val625 630 635 640Met
Lys Ile Ala Asp Phe Gly Leu Ala Arg Asp Ile Asn Asn Ile Asp 645 650
655Tyr Tyr Lys Lys Thr Thr Asn Gly Arg Leu Pro Val Lys Trp Met Ala
660 665 670Pro Glu Ala Leu Phe Asp Arg Val Tyr Thr His Gln Ser Asp
Val Trp 675 680 685Ser Phe Gly Val Leu Met Trp Glu Ile Phe Thr Leu
Gly Gly Ser Pro 690 695 700Tyr Pro Gly Ile Pro Val Glu Glu Leu Phe
Lys Leu Leu Lys Glu Gly705 710 715 720His Arg Met Asp Lys Pro Ala
Asn Cys Thr Asn Glu Leu Tyr Met Met 725 730 735Met Arg Asp Cys Trp
His Ala Val Pro Ser Gln Arg Pro Thr Phe Lys 740 745 750Gln Leu Val
Glu Asp Leu Asp Arg Ile Leu Thr Leu Thr Thr Asn Glu 755 760 765Glu
Tyr Leu Asp Leu Ser Gln Pro Leu Glu Gln Tyr Ser Pro Ser Tyr 770 775
780Pro Asp Thr Arg Ser Ser Cys Ser Ser Gly Asp Asp Ser Val Phe
Ser785 790 795 800Pro Asp Pro Met Pro Tyr Glu Pro Cys Leu Pro Gln
Tyr Pro His Ile 805 810 815Asn Gly Ser Val Lys Thr 8205806PRTHomo
sapien 5Met Gly Ala Pro Ala Cys Ala Leu Ala Leu Cys Val Ala Val Ala
Ile1 5 10 15Val Ala Gly Ala Ser Ser Glu Ser Leu Gly Thr Glu Gln Arg
Val Val 20 25 30Gly Arg Ala Ala Glu Val Pro Gly Pro Glu Pro Gly Gln
Gln Glu Gln 35 40 45Leu Val Phe Gly Ser Gly Asp Ala Val Glu Leu Ser
Cys Pro Pro Pro 50 55 60Gly Gly Gly Pro Met Gly Pro Thr Val Trp Val
Lys Asp Gly Thr Gly65 70 75 80Leu Val Pro Ser Glu Arg Val Leu Val
Gly Pro Gln Arg Leu Gln Val 85 90 95Leu Asn Ala Ser His Glu Asp Ser
Gly Ala Tyr Ser Cys Arg Gln Arg 100 105 110Leu Thr Gln Arg Val Leu
Cys His Phe Ser Val Arg Val Thr Asp Ala 115 120 125Pro Ser Ser Gly
Asp Asp Glu Asp Gly Glu Asp Glu Ala Glu Asp Thr 130 135 140Gly Val
Asp Thr Gly Ala Pro Tyr Trp Thr Arg Pro Glu Arg Met Asp145 150 155
160Lys Lys Leu Leu Ala Val Pro Ala Ala Asn Thr Val Arg Phe Arg Cys
165 170 175Pro Ala Ala Gly Asn Pro Thr Pro Ser Ile Ser Trp Leu Lys
Asn Gly 180 185 190Arg Glu Phe Arg Gly Glu His Arg Ile Gly Gly Ile
Lys Leu Arg His 195 200 205Gln Gln Trp Ser Leu Val Met Glu Ser Val
Val Pro Ser Asp Arg Gly 210 215 220Asn Tyr Thr Cys Val Val Glu Asn
Lys Phe Gly Ser Ile Arg Gln Thr225 230 235 240Tyr Thr Leu Asp Val
Leu Glu Arg Ser Pro His Arg Pro Ile Leu Gln 245 250 255Ala Gly Leu
Pro Ala Asn Gln Thr Ala Val Leu Gly Ser Asp Val Glu 260 265 270Phe
His Cys Lys Val Tyr Ser Asp Ala Gln Pro His Ile Gln Trp Leu 275 280
285Lys His Val Glu Val Asn Gly Ser Lys Val Gly Pro Asp Gly Thr Pro
290 295 300Tyr Val Thr Val Leu Lys Thr Ala Gly Ala Asn Thr Thr Asp
Lys Glu305 310 315 320Leu Glu Val Leu Ser Leu His Asn Val Thr Phe
Glu Asp Ala Gly Glu 325 330 335Tyr Thr Cys Leu Ala Gly Asn Ser Ile
Gly Phe Ser His His Ser Ala 340 345 350Trp Leu Val Val Leu Pro Ala
Glu Glu Glu Leu Val Glu Ala Asp Glu 355 360 365Ala Gly Ser Val Tyr
Ala Gly Ile Leu Ser Tyr Gly Val Gly Phe Phe 370 375 380Leu Phe Ile
Leu Val Val Ala Ala Val Thr Leu Cys Arg Leu Arg Ser385 390 395
400Pro Pro Lys Lys Gly Leu Gly Ser Pro Thr Val His Lys Ile Ser Arg
405 410 415Phe Pro Leu Lys Arg Gln Val Ser Leu Glu Ser Asn Ala Ser
Met Ser 420 425 430Ser Asn Thr Pro Leu Val Arg Ile Ala Arg Leu Ser
Ser Gly Glu Gly 435 440 445Pro Thr Leu Ala Asn Val Ser Glu Leu Glu
Leu Pro Ala Asp Pro Lys 450 455 460Trp Glu Leu Ser Arg Ala Arg Leu
Thr Leu Gly Lys Pro Leu Gly Glu465 470 475 480Gly Cys Phe Gly Gln
Val Val Met Ala Glu Ala Ile Gly Ile Asp Lys 485 490 495Asp Arg Ala
Ala Lys Pro Val Thr Val Ala Val Lys Met Leu Lys Asp 500 505 510Asp
Ala Thr Asp Lys Asp Leu Ser Asp Leu Val Ser Glu Met Glu Met 515 520
525Met Lys Met Ile Gly Lys His Lys Asn Ile Ile Asn Leu Leu Gly Ala
530 535 540Cys Thr Gln Gly Gly Pro Leu Tyr Val Leu Val Glu Tyr Ala
Ala Lys545 550 555 560Gly Asn Leu Arg Glu Phe Leu Arg Ala Arg Arg
Pro Pro Gly Leu Asp 565 570 575Tyr Ser Phe Asp Thr Cys Lys Pro Pro
Glu Glu Gln Leu Thr Phe Lys 580 585 590Asp Leu Val Ser Cys Ala Tyr
Gln Val Ala Arg Gly Met Glu Tyr Leu 595 600 605Ala Ser Gln Lys Cys
Ile His Arg Asp Leu Ala Ala Arg Asn Val Leu 610 615 620Val Thr Glu
Asp Asn Val Met Lys Ile Ala Asp Phe Gly Leu Ala Arg625 630 635
640Asp Val His Asn Leu Asp Tyr Tyr Lys Lys Thr Thr Asn Gly Arg Leu
645 650 655Pro Val Lys Trp Met Ala Pro Glu Ala Leu Phe Asp Arg Val
Tyr Thr 660 665 670His Gln Ser Asp Val Trp Ser Phe Gly Val Leu Leu
Trp Glu Ile Phe 675 680 685Thr Leu Gly Gly Ser Pro Tyr Pro Gly Ile
Pro Val Glu Glu Leu Phe 690 695 700Lys Leu Leu Lys Glu Gly His Arg
Met Asp Lys Pro Ala Asn Cys Thr705 710 715 720His Asp Leu Tyr Met
Ile Met Arg Glu Cys Trp His Ala Ala Pro Ser 725 730 735Gln Arg Pro
Thr Phe Lys Gln Leu Val Glu Asp Leu Asp Arg Val Leu 740 745 750Thr
Val Thr Ser Thr Asp Glu Tyr Leu Asp Leu Ser Ala Pro Phe Glu 755 760
765Gln Tyr Ser Pro Gly Gly Gln Asp Thr Pro Ser Ser Ser Ser Ser Gly
770 775 780Asp Asp Ser Val Phe Ala His Asp Leu Leu Pro Pro Ala Pro
Pro Ser785 790 795 800Ser Gly Gly Ser Arg Thr 8056808PRTHomo
sapiens 6Met Gly Ala Pro Ala Cys Ala Leu Ala Leu Cys Val Ala Val
Ala Ile1 5 10 15Val Ala Gly Ala Ser Ser Glu Ser Leu Gly Thr Glu Gln
Arg Val Val 20 25 30Gly Arg Ala Ala Glu Val Pro Gly Pro Glu Pro Gly
Gln Gln Glu Gln 35 40 45Leu Val Phe Gly Ser Gly Asp Ala Val Glu Leu
Ser Cys Pro Pro Pro 50 55 60Gly Gly Gly Pro Met Gly Pro Thr Val Trp
Val Lys Asp Gly Thr Gly65 70 75 80Leu Val Pro Ser Glu Arg Val Leu
Val Gly Pro Gln Arg Leu Gln Val 85 90 95Leu Asn Ala Ser His Glu Asp
Ser Gly Ala Tyr Ser Cys Arg Gln Arg 100 105 110Leu Thr Gln Arg Val
Leu Cys His Phe Ser Val Arg Val Thr Asp Ala 115 120 125Pro Ser Ser
Gly Asp Asp Glu Asp Gly Glu Asp Glu Ala Glu Asp Thr 130 135 140Gly
Val Asp Thr Gly Ala Pro Tyr Trp Thr Arg Pro Glu Arg Met Asp145 150
155 160Lys Lys Leu Leu Ala Val Pro Ala Ala Asn Thr Val Arg Phe Arg
Cys 165 170 175Pro Ala Ala Gly Asn Pro Thr Pro Ser Ile Ser Trp Leu
Lys Asn Gly 180 185 190Arg Glu Phe Arg Gly Glu His Arg Ile Gly Gly
Ile Lys Leu Arg His 195 200 205Gln Gln Trp Ser Leu Val Met Glu Ser
Val Val Pro Ser Asp Arg Gly 210 215 220Asn Tyr Thr Cys Val Val Glu
Asn Lys Phe Gly Ser Ile Arg Gln Thr225 230 235 240Tyr Thr Leu Asp
Val Leu Glu Arg Ser Pro His Arg Pro Ile Leu Gln 245 250 255Ala Gly
Leu Pro Ala Asn Gln Thr Ala Val Leu Gly Ser Asp Val Glu 260 265
270Phe His Cys Lys Val Tyr Ser Asp Ala Gln Pro His Ile Gln Trp Leu
275 280 285Lys His Val Glu Val Asn Gly Ser Lys Val Gly Pro Asp Gly
Thr Pro 290 295 300Tyr Val Thr Val Leu Lys Ser Trp Ile Ser Glu Ser
Val Glu Ala Asp305 310 315 320Val Arg Leu Arg Leu Ala Asn Val Ser
Glu Arg Asp Gly Gly Glu Tyr 325 330 335Leu Cys Arg Ala Thr Asn Phe
Ile Gly Val Ala Glu Lys Ala Phe Trp 340 345 350Leu Ser Val His Gly
Pro Arg Ala Ala Glu Glu Glu Leu Val Glu Ala 355 360 365Asp Glu Ala
Gly Ser Val Tyr Ala Gly Ile Leu Ser Tyr Gly Val Gly 370 375 380Phe
Phe Leu Phe Ile Leu Val Val Ala Ala Val Thr Leu Cys Arg Leu385 390
395 400Arg Ser Pro Pro Lys Lys Gly Leu Gly Ser Pro Thr Val His Lys
Ile 405 410 415Ser Arg Phe Pro Leu Lys Arg Gln Val Ser Leu Glu Ser
Asn Ala Ser 420 425 430Met Ser Ser Asn Thr Pro Leu Val Arg Ile Ala
Arg Leu Ser Ser Gly 435 440 445Glu Gly Pro Thr Leu Ala Asn Val Ser
Glu Leu Glu Leu Pro Ala Asp 450 455 460Pro Lys Trp Glu Leu Ser Arg
Ala Arg Leu Thr Leu Gly Lys Pro Leu465 470 475 480Gly Glu Gly Cys
Phe Gly Gln Val Val Met Ala Glu Ala Ile Gly Ile 485 490 495Asp Lys
Asp Arg Ala Ala Lys Pro Val Thr Val Ala Val Lys Met Leu 500 505
510Lys Asp Asp Ala Thr Asp Lys Asp Leu Ser Asp Leu Val Ser Glu Met
515 520 525Glu Met Met Lys Met Ile Gly Lys His Lys Asn Ile Ile Asn
Leu Leu 530 535 540Gly Ala Cys Thr Gln Gly Gly Pro Leu Tyr Val Leu
Val Glu Tyr Ala545 550 555 560Ala Lys Gly Asn Leu Arg Glu Phe Leu
Arg Ala Arg Arg Pro Pro Gly 565 570 575Leu Asp Tyr Ser Phe Asp Thr
Cys Lys Pro Pro Glu Glu Gln Leu Thr 580 585 590Phe Lys Asp Leu Val
Ser Cys Ala Tyr Gln Val Ala Arg Gly Met Glu 595 600 605Tyr Leu Ala
Ser Gln Lys Cys Ile His Arg Asp Leu Ala Ala Arg Asn 610 615 620Val
Leu Val Thr Glu Asp Asn Val Met Lys Ile Ala Asp Phe Gly Leu625 630
635 640Ala Arg Asp Val His Asn Leu Asp Tyr Tyr Lys Lys Thr Thr Asn
Gly 645 650 655Arg Leu Pro Val Lys Trp Met Ala Pro Glu Ala Leu Phe
Asp Arg Val 660 665 670Tyr Thr His Gln Ser Asp Val Trp Ser Phe Gly
Val Leu Leu Trp Glu 675 680 685Ile Phe Thr Leu Gly Gly Ser Pro Tyr
Pro Gly Ile Pro Val Glu Glu 690 695 700Leu Phe Lys Leu Leu Lys Glu
Gly His Arg Met Asp Lys Pro Ala Asn705 710 715 720Cys Thr His Asp
Leu Tyr Met Ile Met Arg Glu Cys Trp His Ala Ala 725 730 735Pro Ser
Gln Arg Pro Thr Phe Lys Gln Leu Val Glu Asp Leu Asp Arg 740 745
750Val Leu Thr Val Thr Ser Thr Asp Glu Tyr Leu Asp Leu Ser Ala Pro
755 760 765Phe Glu Gln Tyr Ser Pro Gly Gly Gln Asp Thr Pro Ser Ser
Ser Ser 770 775 780Ser Gly Asp Asp Ser Val Phe Ala His Asp Leu Leu
Pro Pro Ala Pro785 790 795 800Pro Ser Ser Gly Gly Ser Arg Thr
8057802PRTHomo sapiens 7Met Arg Leu Leu Leu Ala Leu Leu Gly Val Leu
Leu Ser Val Pro Gly1 5 10 15Pro Pro Val Leu Ser Leu Glu Ala Ser Glu
Glu Val Glu Leu Glu Pro 20 25 30Cys Leu
Ala Pro Ser Leu Glu Gln Gln Glu Gln Glu Leu Thr Val Ala 35 40 45Leu
Gly Gln Pro Val Arg Leu Cys Cys Gly Arg Ala Glu Arg Gly Gly 50 55
60His Trp Tyr Lys Glu Gly Ser Arg Leu Ala Pro Ala Gly Arg Val Arg65
70 75 80Gly Trp Arg Gly Arg Leu Glu Ile Ala Ser Phe Leu Pro Glu Asp
Ala 85 90 95Gly Arg Tyr Leu Cys Leu Ala Arg Gly Ser Met Ile Val Leu
Gln Asn 100 105 110Leu Thr Leu Ile Thr Gly Asp Ser Leu Thr Ser Ser
Asn Asp Asp Glu 115 120 125Asp Pro Lys Ser His Arg Asp Pro Ser Asn
Arg His Ser Tyr Pro Gln 130 135 140Gln Ala Pro Tyr Trp Thr His Pro
Gln Arg Met Glu Lys Lys Leu His145 150 155 160Ala Val Pro Ala Gly
Asn Thr Val Lys Phe Arg Cys Pro Ala Ala Gly 165 170 175Asn Pro Thr
Pro Thr Ile Arg Trp Leu Lys Asp Gly Gln Ala Phe His 180 185 190Gly
Glu Asn Arg Ile Gly Gly Ile Arg Leu Arg His Gln His Trp Ser 195 200
205Leu Val Met Glu Ser Val Val Pro Ser Asp Arg Gly Thr Tyr Thr Cys
210 215 220Leu Val Glu Asn Ala Val Gly Ser Ile Arg Tyr Asn Tyr Leu
Leu Asp225 230 235 240Val Leu Glu Arg Ser Pro His Arg Pro Ile Leu
Gln Ala Gly Leu Pro 245 250 255Ala Asn Thr Thr Ala Val Val Gly Ser
Asp Val Glu Leu Leu Cys Lys 260 265 270Val Tyr Ser Asp Ala Gln Pro
His Ile Gln Trp Leu Lys His Ile Val 275 280 285Ile Asn Gly Ser Ser
Phe Gly Ala Asp Gly Phe Pro Tyr Val Gln Val 290 295 300Leu Lys Thr
Ala Asp Ile Asn Ser Ser Glu Val Glu Val Leu Tyr Leu305 310 315
320Arg Asn Val Ser Ala Glu Asp Ala Gly Glu Tyr Thr Cys Leu Ala Gly
325 330 335Asn Ser Ile Gly Leu Ser Tyr Gln Ser Ala Trp Leu Thr Val
Leu Pro 340 345 350Glu Glu Asp Pro Thr Trp Thr Ala Ala Ala Pro Glu
Ala Arg Tyr Thr 355 360 365Asp Ile Ile Leu Tyr Ala Ser Gly Ser Leu
Ala Leu Ala Val Leu Leu 370 375 380Leu Leu Ala Gly Leu Tyr Arg Gly
Gln Ala Leu His Gly Arg His Pro385 390 395 400Arg Pro Pro Ala Thr
Val Gln Lys Leu Ser Arg Phe Pro Leu Ala Arg 405 410 415Gln Phe Ser
Leu Glu Ser Gly Ser Ser Gly Lys Ser Ser Ser Ser Leu 420 425 430Val
Arg Gly Val Arg Leu Ser Ser Ser Gly Pro Ala Leu Leu Ala Gly 435 440
445Leu Val Ser Leu Asp Leu Pro Leu Asp Pro Leu Trp Glu Phe Pro Arg
450 455 460Asp Arg Leu Val Leu Gly Lys Pro Leu Gly Glu Gly Cys Phe
Gly Gln465 470 475 480Val Val Arg Ala Glu Ala Phe Gly Met Asp Pro
Ala Arg Pro Asp Gln 485 490 495Ala Ser Thr Val Ala Val Lys Met Leu
Lys Asp Asn Ala Ser Asp Lys 500 505 510Asp Leu Ala Asp Leu Val Ser
Glu Met Glu Val Met Lys Leu Ile Gly 515 520 525Arg His Lys Asn Ile
Ile Asn Leu Leu Gly Val Cys Thr Gln Glu Gly 530 535 540Pro Leu Tyr
Val Ile Val Glu Cys Ala Ala Lys Gly Asn Leu Arg Glu545 550 555
560Phe Leu Arg Ala Arg Arg Pro Pro Gly Pro Asp Leu Ser Pro Asp Gly
565 570 575Pro Arg Ser Ser Glu Gly Pro Leu Ser Phe Pro Val Leu Val
Ser Cys 580 585 590Ala Tyr Gln Val Ala Arg Gly Met Gln Tyr Leu Glu
Ser Arg Lys Cys 595 600 605Ile His Arg Asp Leu Ala Ala Arg Asn Val
Leu Val Thr Glu Asp Asn 610 615 620Val Met Lys Ile Ala Asp Phe Gly
Leu Ala Arg Gly Val His His Ile625 630 635 640Asp Tyr Tyr Lys Lys
Thr Ser Asn Gly Arg Leu Pro Val Lys Trp Met 645 650 655Ala Pro Glu
Ala Leu Phe Asp Arg Val Tyr Thr His Gln Ser Asp Val 660 665 670Trp
Ser Phe Gly Ile Leu Leu Trp Glu Ile Phe Thr Leu Gly Gly Ser 675 680
685Pro Tyr Pro Gly Ile Pro Val Glu Glu Leu Phe Ser Leu Leu Arg Glu
690 695 700Gly His Arg Met Asp Arg Pro Pro His Cys Pro Pro Glu Leu
Tyr Gly705 710 715 720Leu Met Arg Glu Cys Trp His Ala Ala Pro Ser
Gln Arg Pro Thr Phe 725 730 735Lys Gln Leu Val Glu Ala Leu Asp Lys
Val Leu Leu Ala Val Ser Glu 740 745 750Glu Tyr Leu Asp Leu Arg Leu
Thr Phe Gly Pro Tyr Ser Pro Ser Gly 755 760 765Gly Asp Ala Ser Ser
Thr Cys Ser Ser Ser Asp Ser Val Phe Ser His 770 775 780Asp Pro Leu
Pro Leu Gly Ser Ser Ser Phe Pro Phe Gly Ser Gly Val785 790 795
800Gln Thr8762PRTHomo sapiens 8Met Arg Leu Leu Leu Ala Leu Leu Gly
Val Leu Leu Ser Val Pro Gly1 5 10 15Pro Pro Val Leu Ser Leu Glu Ala
Ser Glu Glu Val Glu Leu Glu Pro 20 25 30Cys Leu Ala Pro Ser Leu Glu
Gln Gln Glu Gln Glu Leu Thr Val Ala 35 40 45Leu Gly Gln Pro Val Arg
Leu Cys Cys Gly Arg Ala Glu Arg Gly Gly 50 55 60His Trp Tyr Lys Glu
Gly Ser Arg Leu Ala Pro Ala Gly Arg Val Arg65 70 75 80Gly Trp Arg
Gly Arg Leu Glu Ile Ala Ser Phe Leu Pro Glu Asp Ala 85 90 95Gly Arg
Tyr Leu Cys Leu Ala Arg Gly Ser Met Ile Val Leu Gln Asn 100 105
110Leu Thr Leu Ile Thr Gly Asp Ser Leu Thr Ser Ser Asn Asp Asp Glu
115 120 125Asp Pro Lys Ser His Arg Asp Pro Ser Asn Arg His Ser Tyr
Pro Gln 130 135 140Gln Ala Pro Tyr Trp Thr His Pro Gln Arg Met Glu
Lys Lys Leu His145 150 155 160Ala Val Pro Ala Gly Asn Thr Val Lys
Phe Arg Cys Pro Ala Ala Gly 165 170 175Asn Pro Thr Pro Thr Ile Arg
Trp Leu Lys Asp Gly Gln Ala Phe His 180 185 190Gly Glu Asn Arg Ile
Gly Gly Ile Arg Leu Arg His Gln His Trp Ser 195 200 205Leu Val Met
Glu Ser Val Val Pro Ser Asp Arg Gly Thr Tyr Thr Cys 210 215 220Leu
Val Glu Asn Ala Val Gly Ser Ile Arg Tyr Asn Tyr Leu Leu Asp225 230
235 240Val Leu Glu Arg Ser Pro His Arg Pro Ile Leu Gln Ala Gly Leu
Pro 245 250 255Ala Asn Thr Thr Ala Val Val Gly Ser Asp Val Glu Leu
Leu Cys Lys 260 265 270Val Tyr Ser Asp Ala Gln Pro His Ile Gln Trp
Leu Lys His Ile Val 275 280 285Ile Asn Gly Ser Ser Phe Gly Ala Asp
Gly Phe Pro Tyr Val Gln Val 290 295 300Leu Lys Thr Ala Asp Ile Asn
Ser Ser Glu Val Glu Val Leu Tyr Leu305 310 315 320Arg Asn Val Ser
Ala Glu Asp Ala Gly Glu Tyr Thr Cys Leu Ala Gly 325 330 335Asn Ser
Ile Gly Leu Ser Tyr Gln Ser Ala Trp Leu Thr Val Leu Pro 340 345
350Gly Thr Gly Arg Ile Pro His Leu Thr Cys Asp Ser Leu Thr Pro Ala
355 360 365Gly Arg Thr Lys Ser Pro Thr Leu Gln Phe Ser Leu Glu Ser
Gly Ser 370 375 380Ser Gly Lys Ser Ser Ser Ser Leu Val Arg Gly Val
Arg Leu Ser Ser385 390 395 400Ser Gly Pro Ala Leu Leu Ala Gly Leu
Val Ser Leu Asp Leu Pro Leu 405 410 415Asp Pro Leu Trp Glu Phe Pro
Arg Asp Arg Leu Val Leu Gly Lys Pro 420 425 430Leu Gly Glu Gly Cys
Phe Gly Gln Val Val Arg Ala Glu Ala Phe Gly 435 440 445Met Asp Pro
Ala Arg Pro Asp Gln Ala Ser Thr Val Ala Val Lys Met 450 455 460Leu
Lys Asp Asn Ala Ser Asp Lys Asp Leu Ala Asp Leu Val Ser Glu465 470
475 480Met Glu Val Met Lys Leu Ile Gly Arg His Lys Asn Ile Ile Asn
Leu 485 490 495Leu Gly Val Cys Thr Gln Glu Gly Pro Leu Tyr Val Ile
Val Glu Cys 500 505 510Ala Ala Lys Gly Asn Leu Arg Glu Phe Leu Arg
Ala Arg Arg Pro Pro 515 520 525Gly Pro Asp Leu Ser Pro Asp Gly Pro
Arg Ser Ser Glu Gly Pro Leu 530 535 540Ser Phe Pro Val Leu Val Ser
Cys Ala Tyr Gln Val Ala Arg Gly Met545 550 555 560Gln Tyr Leu Glu
Ser Arg Lys Cys Ile His Arg Asp Leu Ala Ala Arg 565 570 575Asn Val
Leu Val Thr Glu Asp Asn Val Met Lys Ile Ala Asp Phe Gly 580 585
590Leu Ala Arg Gly Val His His Ile Asp Tyr Tyr Lys Lys Thr Ser Asn
595 600 605Gly Arg Leu Pro Val Lys Trp Met Ala Pro Glu Ala Leu Phe
Asp Arg 610 615 620Val Tyr Thr His Gln Ser Asp Val Trp Ser Phe Gly
Ile Leu Leu Trp625 630 635 640Glu Ile Phe Thr Leu Gly Gly Ser Pro
Tyr Pro Gly Ile Pro Val Glu 645 650 655Glu Leu Phe Ser Leu Leu Arg
Glu Gly His Arg Met Asp Arg Pro Pro 660 665 670His Cys Pro Pro Glu
Leu Tyr Gly Leu Met Arg Glu Cys Trp His Ala 675 680 685Ala Pro Ser
Gln Arg Pro Thr Phe Lys Gln Leu Val Glu Ala Leu Asp 690 695 700Lys
Val Leu Leu Ala Val Ser Glu Glu Tyr Leu Asp Leu Arg Leu Thr705 710
715 720Phe Gly Pro Tyr Ser Pro Ser Gly Gly Asp Ala Ser Ser Thr Cys
Ser 725 730 735Ser Ser Asp Ser Val Phe Ser His Asp Pro Leu Pro Leu
Gly Ser Ser 740 745 750Ser Phe Pro Phe Gly Ser Gly Val Gln Thr 755
760
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